Performance information

• Visualizing thiazolidine ring formation in the reaction of D-cysteine and pyridoxal-5′-phosphate within L-cysteine desulfurase SufS　　　　　　　　　　　　　　　
  Ryosuke Nakamura; Takashi Fujishiro
  Biochemical and Biophysical Research Communications, Volume：754, First page：151497, Last page：151497, Mar. 2025, [Reviewed], [Last, Corresponding]
  Elsevier BV, English, Scientific journal
  DOI：https://doi.org/10.1016/j.bbrc.2025.151497
  DOI ID：10.1016/j.bbrc.2025.151497, ISSN：0006-291X, 共同研究・競争的資金等ID：42101093
• Structural insights into the recognition of tetrapyrrole substrates by ancestral class II chelatase CfbA
  Shoko Ogawa; Masahide Hikita; Takashi Fujishiro
  Protein Science, Volume：33, Number：12, First page：e5208, Nov. 2024, [Reviewed], [Last, Corresponding]
  Abstract
  
  Nickel‐chelatase CfbA, unlike descendant chelatases, is an ancestral class II chelatase with a symmetric active site architecture. CfbA utilizes sirohydrochlorin (SHC) as a physiological substrate in the biosynthesis of coenzyme F430. CbiX^S, a structural analog of CfbA, can use uroporphyrin III (UPIII) and uroporphyrin I (UPI) as non‐physiological substrates. Owing to the broad tetrapyrrole specificity of the unique active site of ancestral class II chelatases, the substrate recognition mechanism of CfbA has garnered interest. Herein, we conducted an X‐ray crystallographic analysis of CfbA in complex with UPIII and UPI. Interestingly, the binding sites for UPIII and UPI were distinct. UPI was bound at the entrance of the active site, whereas UPIII was bound deep inside the active site cavity in a manner similar to SHC. Despite the difference in the binding positions of UPIII and UPI, Ser11 at the active site provided critical polar interactions for recognizing UPIII and UPI. Several CfbA variants with a Ser11 mutation were studied to confirm the significance of Ser11's position in the context of tetrapyrrole recognition. The CfbA S11T variant showed Ni²⁺‐chelatase activity against coproporphyrin I (CPI), which is a more hydrophobic tetrapyrrole than UPIII and UPI. Using a CPI‐docked model of the S11T variant, we proposed that balancing the hydrophobic/polar interactions at residue 11 could alter substrate selectivity. The structural and mutational analyses reported here highlight the importance of polar and hydrophobic interactions at the entry region of the active site for substrate tetrapyrrole recognition by ancestral and descendant class II chelatases.
  Wiley, English, Scientific journal
  DOI：https://doi.org/10.1002/pro.5208
  DOI ID：10.1002/pro.5208, ISSN：0961-8368, eISSN：1469-896X, ORCID：171853481, 共同研究・競争的資金等ID：42101093;25988841
• Cysteine-Persulfide Sulfane Sulfur-Ligated Zn Complex of Sulfur-Carrying SufU in the SufCDSUB System for Fe–S Cluster Biosynthesis　　　　　　　　　　　　　　　
  Takuya Terahata; Yukino Shimada; Chisato Maki; Suguru Muroga; Rina Sakurai; Kouhei Kunichika; Takashi Fujishiro
  Inorganic Chemistry, Volume：63, Number：42, First page：19607, Last page：19618, Oct. 2024, [Reviewed], [Last, Corresponding]
  English, Scientific journal
  DOI：https://doi.org/10.1021/acs.inorgchem.4c02654
  DOI ID：10.1021/acs.inorgchem.4c02654, ORCID：169287095, 共同研究・競争的資金等ID：42101093;42101092;12335178
• Nickel-chelatase activity of SirB variants mimicking the His arrangement in the naturally occurring nickel-chelatase CfbA.　　　　　　　　　　　　　　　
  Yuuma Oyamada; Shoko Ogawa; Takashi Fujishiro
  FEBS Open Bio, Volume：14, Number：8, First page：1291, Last page：1302, Jun. 2024, [Reviewed], [Last, Corresponding], ［International magazine］
  Metal-tetrapyrrole cofactors are involved in multiple cellular functions, and chelatases are key enzymes for the biosynthesis of these cofactors. CfbA is an ancestral, homodimeric-type class II chelatase which is able to use not only Ni2+ as a physiological metal substrate, but also Co2+ as a nonphysiological substrate with higher activity than for Ni2+. The Ni/Co-chelatase function found in CfbA is also observed in SirB, a descendant, monomeric-type class II chelatase. This is despite the distinct active site structure of CfbA and SirB; specifically, CfbA shows a unique four His residue arrangement, unlike other monomeric class II chelatases such as SirB. Herein, we studied the Ni-chelatase activity of SirB variants R134H, L200H, and R134H/L200H, the latter of which mimics the His alignment of CfbA. Our results showed that the SirB R134H variant exhibited the highest Ni-chelatase activity among the SirB enzymes, which in turn suggests that the position of His134 could be more important for the Ni-chelatase activity than that of His200. The SirB R134H/L200H variant showed lower activity than R134H, despite the four His residues found in SirB R134H/L200H. CD spectroscopy showed secondary structure denaturation and a slight difficulty in Ni-binding of SirB R134H/L200H, which may be related to its lower activity. Finally, a docking simulation suggested that the His134 of the SirB R134H variant could function as a base catalyst for the Ni-chelatase reaction in a class II chelatase architecture.
  English, Scientific journal
  DOI：https://doi.org/10.1002/2211-5463.13849
  DOI ID：10.1002/2211-5463.13849, PubMed ID：38923868, 共同研究・競争的資金等ID：42101093;25988841
• Phage single-stranded DNA-binding protein or host DNA damage triggers the activation of the AbpAB phage defense system　　　　　　　　　　　　　　　
  Takaomi Sasaki; Saya Takita; Takashi Fujishiro; Yunosuke Shintani; Satoki Nojiri; Ryota Yasui; Tetsuro Yonesaki; Yuichi Otsuka
  mSphere, Volume：8, Number：6, First page：e0037223, Dec. 2023, [Reviewed]
  ABSTRACT
  
  Bacteria have developed various defense mechanisms against phages. Abortive infection (Abi), a bacterial defense mechanism, can be achieved through various means, including toxin-antitoxin systems, cyclic oligonucleotide-based antiphage signaling systems, and retrons. AbpA and AbpB (AbpAB) defend against many lytic phages harboring double-stranded DNA genomes in Escherichia coli ; however, how AbpAB senses phage infection and inhibits its propagation remains unclear. Here, we demonstrated that AbpAB inhibited the growth of the φX174 lytic phage with single-stranded DNA (ssDNA) as well as the lysogenization and induction of the Sakai prophage 5 lysogenic phage. The AbpAB defense system limits T4 and φX174 phage propagation via Abi. AbpA contains a nuclease domain at its N-terminus, and AbpB has an ATP-dependent RNA helicase domain; both domains are required for phage defense. This system is activated by phage Gp32 binding to ssDNA and inhibits E. coli growth. Without phage infection, DNA replication inhibitors or defects in the DNA repair factors RecB and RecC activate this system. Therefore, the E. coli AbpAB defense system may sense DNA-protein complexes, including the phage-encoded ssDNA-binding protein or those formed by interrupting host DNA replication or repair.
  
  IMPORTANCE
  
  Although numerous phage defense systems have recently been discovered in bacteria, how these systems defend against phage propagation or sense phage infections remains unclear. The Escherichia coli AbpAB defense system targets several lytic and lysogenic phages harboring DNA genomes. A phage-encoded single-stranded DNA-binding protein, Gp32, activates this system similar to other phage defense systems such as Retron-Eco8, Hachiman, ShosTA, Nhi, and Hna. DNA replication inhibitors or defects in DNA repair factors activate the AbpAB system, even without phage infection. This is one of the few examples of activating phage defense systems without phage infection or proteins. The AbpAB defense system may be activated by sensing specific DNA-protein complexes.
  American Society for Microbiology, English, Scientific journal
  DOI：https://doi.org/10.1128/msphere.00372-23
  DOI ID：10.1128/msphere.00372-23, eISSN：2379-5042, PubMed ID：37882551, PubMed Central ID：PMC10732053
• Class III hybrid cluster protein homodimeric architecture shows evolutionary relationship with Ni, Fe-carbon monoxide dehydrogenases
  Takashi Fujishiro; Kyosei Takaoka
  Nature Communications, Volume：14, Number：1, First page：5609, Sep. 2023, [Reviewed], [Lead, Corresponding]
  Abstract
  
  Hybrid cluster proteins (HCPs) are Fe-S-O cluster-containing metalloenzymes in three distinct classes (class I and II: monomer, III: homodimer), all of which structurally related to homodimeric Ni, Fe-carbon monoxide dehydrogenases (CODHs). Here we show X-ray crystal structure of class III HCP from Methanothermobacter marburgensis (Mm HCP), demonstrating its homodimeric architecture structurally resembles those of CODHs. Also, despite the different architectures of class III and I/II HCPs, [4Fe-4S] and hybrid clusters are found in equivalent positions in all HCPs. Structural comparison of Mm HCP and CODHs unveils some distinct features such as the environments of their homodimeric interfaces and the active site metalloclusters. Furthermore, structural analysis of Mm HCP C67Y and characterization of several Mm HCP variants with a Cys67 mutation reveal the significance of Cys67 in protein structure, metallocluster binding and hydroxylamine reductase activity. Structure-based bioinformatics analysis of HCPs and CODHs provides insights into the structural evolution of the HCP/CODH superfamily.
  Springer Science and Business Media LLC, Scientific journal
  DOI：https://doi.org/10.1038/s41467-023-41289-4
  DOI ID：10.1038/s41467-023-41289-4, eISSN：2041-1723, 共同研究・競争的資金等ID：42101092;37071571
• Escherichia coli Class II Hybrid Cluster Protein, HCP
  Takashi Fujishiro
  Encyclopedia of Inorganic and Bioinorganic Chemistry, First page：1, Last page：7, Apr. 2023, [Reviewed], [Lead, Last, Corresponding]
  Abstract
  
  Escherichia coli hybrid cluster protein ( Ec HCP) belongs to class II of the HCP family that harbors an iron–sulfur–oxygen (Fe–S–O) cluster as its active site for the reduction of hydroxylamine to ammonia. Like the other class II HCPs, Ec HCP uniquely exhibits a long N‐terminal Cys‐rich motif compared with the other classes (class I and III) of HCPs and exclusively utilizes HCP reductase (HCR) as an electron‐transfer partner protein. Structural and functional analyses of Ec HCP revealed the characteristic features of class II HCPs, especially the uniqueness of the N‐terminal Cys‐rich motif and its surroundings. Notably, the Ec HCP has an N‐terminal [4Fe–4S] cluster rather than a [2Fe–2S] cluster, which had long been believed to be present. The binding mode of the N‐terminal [4Fe–4S] cluster to class II Ec HCP is similar to that of class I HCPs: the N‐terminal [4Fe–4S] cluster is coordinated by four conserved Cys ligands. In contrast, the N‐terminal loop structure of class II Ec HCP was different from that of class I HCPs. Ec HCP has a protruded loop structure in the N‐terminal region and a hydrophobic environment, whereas class I HCPs do not. This uniqueness of the N‐terminal region of Ec HCP was critical for the electron transfer reaction involving the formation of Ec HCP‐ Ec HCR complex with its suitable orientation for electron transfer.
  Wiley, English
  DOI：https://doi.org/10.1002/9781119951438.eibc2839
  DOI ID：10.1002/9781119951438.eibc2839
• Cycloserine enantiomers inhibit PLP‐dependent cysteine desulfurase SufS via distinct mechanisms　　　　　　　　　　　　　　　
  Ryosuke Nakamura; Shoko Ogawa; Yasuhiro Takahashi; Takashi Fujishiro
  The FEBS Journal, Volume：289, Number：19, First page：5947, Last page：5970, Apr. 2022, [Reviewed], [Last, Corresponding]
  The cysteine desulfurase SufS is a pyridoxal‐5′‐phosphate‐dependent enzyme and is essential for the SUF system, which participates in iron–sulfur cluster biosynthesis. Inhibition of SufS in the SUF system by D‐cycloserine (DCS) in Plasmodium falciparum apicoplast has recently been reported, indicating that SufS could be a target for malaria therapeutics. However, the mechanistic details underlying the inhibition of SufS by DCS have not yet been clarified. Moreover, inhibition of SufS by the other enantiomer, L‐cycloserine (LCS), has not been investigated. Herein, we investigated the structure‐based inhibition mechanisms of SufS by DCS and LCS using Bacillus subtilis SufS, whose catalytic mechanism has been well characterized in comparison to that of the P. falciparum SufS. Surprisingly, DCS‐ and LCS‐mediated inhibitions of SufS occur via distinct mechanisms resulting in pyridoxamine‐5′‐phosphate (PMP) in DCS‐mediated inhibition and PMP‐3‐hydroxyisoxazole adduct (PMP‐isoxazole) in LCS‐mediated inhibition. Biochemical and structural evaluation of SufS variants identified conserved His and Arg residues at the active site as the key determinants of the distinct inhibition mechanisms. The importance of structural elements involved in DCS and LCS‐mediated inhibitions of SufS provides valuable insights for the structure‐based design of new drugs targeting SufS.
  
  Database
  
  Structural data are available in PDB database under the accession numbers 6KFY, 7CEO, 7CEP, 7CEQ, 7CER, 7CES, 7CET, 7CEU, 7E6A, 7E6B, 7E6C, 7E6D, 7E6E, and 7E6F.
  Wiley, English, Scientific journal
  DOI：https://doi.org/10.1111/febs.16455
  DOI ID：10.1111/febs.16455, ISSN：1742-464X, eISSN：1742-4658, ORCID：130085124, Web of Science ID：WOS:000782913800001
• Sirohydrochlorin Nickelochelatase CfbA
  Takashi Fujishiro
  Encyclopedia of Inorganic and Bioinorganic Chemistry, First page：1, Last page：15, Mar. 2022, [Reviewed], [Lead, Last, Corresponding]
  Wiley, English, Scientific journal
  DOI：https://doi.org/10.1002/9781119951438.eibc2815
  DOI ID：10.1002/9781119951438.eibc2815, ORCID：110346116, 共同研究・競争的資金等ID：25988841
• Structural diversity of cysteine desulfurases involved in iron-sulfur cluster biosynthesis　　　　　　　　　　　　　　　
  Takashi Fujishiro; Ryosuke Nakamura; Kouhei Kunichika; Yasuhiro Takahashi
  Biophysics and Physicobiology, Volume：19, First page：e190001, Last page：1, 2022, [Reviewed], [Last, Corresponding]
  Biophysical Society of Japan, English, Scientific journal
  DOI：https://doi.org/10.2142/biophysico.bppb-v19.0001
  DOI ID：10.2142/biophysico.bppb-v19.0001, ISSN：2189-4779, ORCID：108601522
• Crystal structure of Escherichia coli class II hybrid cluster protein, HCP, reveals a [4Fe-4S] cluster at the N-terminal protrusion.　　　　　　　　　　　　　　　
  Takashi Fujishiro; Miho Ooi; Kyosei Takaoka
  The FEBS journal, Volume：288, Number：23, First page：6752, Last page：6768, Jun. 2021, [Reviewed], [Lead, Corresponding], ［International magazine］
  Hybrid cluster protein (HCP) is a unique Fe-S-O-type metallocluster-containing enzyme present in many anaerobic organisms and is categorized into three distinct classes (I, II, and III). The class II HCP uniquely utilizes hybrid cluster protein reductase (HCR), unlike the other classes of HCPs. To gain structural insights into the electron transfer system between the class II HCP and HCR, we elucidated the X-ray crystal structure of Escherichia coli HCP (Ec HCP), representing the first report of a class II HCP structure. Surprisingly, Ec HCP was found to harbor a [4Fe-4S] cluster rather than a [2Fe-2S] cluster at the N-terminal Cys-rich region, similar to class I HCPs. It was also found that the Cys-rich motif forms a unique protrusion and that the surrounding charge distributions on the surface of class II Ec HCP are distinct from those of class I HCPs. The functional significance of the Cys-rich region was investigated using an Ec HCP variant (chimeric HCP) containing a class I HCP Cys-rich motif from Desulfovibrio desulfuricans. The biochemical analyses showed that the chimeric HCP lacks the hybrid cluster and the electron-accepting function from HCR despite the formation of the chimeric HCP-HCR complex. Furthermore, HCP-HCR molecular docking analysis suggested that the protrusion area serves as an HCR-binding region. Therefore, the protrusion of the unique Cys-rich motif and the surrounding area of class II HCP are likely important for maturation of Ec HCP and orienting HCR onto the surface of HCP to facilitate electron transfer in the HCP-HCR complex.
  English, Scientific journal
  DOI：https://doi.org/10.1111/febs.16062
  DOI ID：10.1111/febs.16062, PubMed ID：34101368
• The Structure of the Dimeric State of IscU Harboring Two Adjacent [2Fe-2S] Clusters Provides Mechanistic Insights into Cluster Conversion to [4Fe-4S].　　　　　　　　　　　　　　　
  Kouhei Kunichika; Ryosuke Nakamura; Takashi Fujishiro; Yasuhiro Takahashi
  Biochemistry, Volume：60, Number：20, First page：1569, Last page：1572, May 2021, [Reviewed], [Corresponding], ［International magazine］
  IscU serves as a scaffold for the de novo assembly of a [2Fe-2S] cluster prior to its delivery to recipient protein. It has also been proposed that on one dimer of bacterial IscU, two [2Fe-2S] clusters can be converted into a single [4Fe-4S] cluster. However, lack of structural information about the dimeric state of IscU has hindered our understanding of the underlying mechanisms. In this study, we determine the X-ray crystal structure of IscU from the thermophilic archaeon Methanothrix thermoacetophila and demonstrate a dimer structure of IscU in which two [2Fe-2S] clusters are facing each other in close proximity at the dimer interface. Our structure also reveals for the first time that Asp40 serves as a fourth ligand to the [2Fe-2S] cluster with three Cys ligands in each monomer, consistent with previous spectroscopic data. We confirm by EPR spectroscopic analysis that in solution two adjacent [2Fe-2S] clusters in the wild-type dimer are converted to a [4Fe-4S] cluster via reductive coupling. Furthermore, we find that the H106A substitution abolishes the reductive conversion to the [4Fe-4S] cluster without structural alteration, suggesting that His106 is functionally involved in this process. Overall, these findings provide a structural explanation for the assembly and conversion of Fe-S clusters on IscU and highlight a dynamic process that advances via association and dissociation of the IscU dimer.
  English, Scientific journal
  DOI：https://doi.org/10.1021/acs.biochem.1c00112
  DOI ID：10.1021/acs.biochem.1c00112, PubMed ID：33938220
• Evidence for dynamic in vivo interconversion of the conformational states of IscU during iron-sulfur cluster biosynthesis.　　　　　　　　　　　　　　　
  Sakiko Sato; Yumeka Matsushima; Miaki Kanazawa; Naoyuki Tanaka; Takashi Fujishiro; Kouhei Kunichika; Ryosuke Nakamura; Hiroaki Tomioka; Kei Wada; Yasuhiro Takahashi
  Molecular microbiology, Volume：115, Number：4, First page：807, Last page：818, Apr. 2021, [Reviewed], ［International magazine］
  IscU is a central component of the ISC machinery and serves as a scaffold for de novo assembly of Fe-S clusters. The dedicated chaperone system composed of the Hsp70-chaperone HscA and the J-protein cochaperone HscB synergistically interacts with IscU and facilitates cluster transfer from IscU to recipient apo-proteins. Here, we report that the otherwise essential roles of HscA and HscB can be bypassed in vivo by a number of single amino acid substitutions in IscU. CD spectroscopic studies of the variant IscU proteins capable of this bypass activity revealed dynamic interconversion between two conformations: the denatured (D) and the structured (S) state in the absence and presence of Zn2+ , respectively, which was far more prominent than interconversion observed in wild-type IscU. Furthermore, we found that neither the S-shifted (more structured) variants of IscU nor the perpetually denatured variants could perform their in vivo role regardless of whether the chaperone system was present or not. The present study thus provides for the first time evidence that an in vivo D-state of IscU exists and implies that conformational interconversion between the S- and D-states of the scaffolding protein is a fundamental requirement for the assembly and transfer of the Fe-S cluster.
  English, Scientific journal
  DOI：https://doi.org/10.1111/mmi.14646
  DOI ID：10.1111/mmi.14646, PubMed ID：33202070
• A cyclic lipopeptide surfactin is a species-selective Hsp90 inhibitor that suppresses cyanobacterial growth.　　　　　　　　　　　　　　　
  Hitoshi Nakamoto; Yuhei Yokoyama; Takahiro Suzuki; Yuri Miyamoto; Takashi Fujishiro; Masaaki Morikawa; Yoshihiko Miyata
  Journal of biochemistry, Mar. 2021, [Reviewed], ［International magazine］
  Heat shock protein 90 (Hsp90) is essential for eukaryotic cells, whereas bacterial homologs play a role under stresses and in pathogenesis. Identifying species-specific Hsp90 inhibitors is challenging because Hsp90 is evolutionarily conserved. We found that a cyclic lipopeptide surfactin inhibits the ATPase activity of Hsp90 from the cyanobacterium Synechococcus elongatus (S. elongatus) PCC 7942 but does not inhibit Escherichia coli (E. coli), yeast and human Hsp90s. Molecular docking simulations indicated that surfactin could bind to the N-terminal dimerization interface of the cyanobacterial Hsp90 in the ATP- and ADP-bound states, which provided molecular insights into the species-selective inhibition. The data suggest that surfactin inhibits a rate-limiting conformational change of S. elongatus Hsp90 in the ATP hydrolysis. Surfactin also inhibited the interaction of the cyanobacterial Hsp90 with a model substrate, and suppressed S. elongatus growth under heat stress, but not that of E. coli. Surfactin did not show significant cellular toxicity toward mammalian cells. These results indicate that surfactin inhibits the cellular function of Hsp90 specifically in the cyanobacterium. The present study shows that a cyclic peptide has a great specificity to interact with a specific homolog of a highly conserved protein family.
  English, Scientific journal
  DOI：https://doi.org/10.1093/jb/mvab037
  DOI ID：10.1093/jb/mvab037, PubMed ID：33768253
• The nickel-sirohydrochlorin formation mechanism of the ancestral class II chelatase CfbA in coenzyme F430 biosynthesis　　　　　　　　　　　　　　　
  Takashi Fujishiro; Shoko Ogawa
  Chemical Science, Volume：12, Number：6, First page：2172, Last page：2180, 2021, [Reviewed], [Lead, Corresponding], ［International magazine］
  

  The substrate-assisted nickel chelatase mechanism of CfbA in coenzyme F430 biosynthesis was unveiled by X-ray crystal structure analysis.

  
  Royal Society of Chemistry (RSC), English, Scientific journal
  DOI：https://doi.org/10.1039/d0sc05439a
  DOI ID：10.1039/d0sc05439a, ISSN：2041-6520, eISSN：2041-6539, PubMed ID：34163982, PubMed Central ID：PMC8179277, 共同研究・競争的資金等ID：25988841
• 2. Hydrogen development
  Ulf-Peter Apfel; Wolfgang Weigand; Marius Horch; Ingo Zebger; Oliver Lenz; Takashi Fujishiro
  Bioorganometallic Chemistry, First page：13, Last page：136, Jun. 2020, [Invited]
  De Gruyter, In book
  DOI：https://doi.org/10.1515/9783110496574-002
  DOI ID：10.1515/9783110496574-002, ORCID：80646571
• Snapshots of PLP‐substrate and PLP‐product external aldimines as intermediates in two types of cysteine desulfurase enzymes　　　　　　　　　　　　　　　
  Ryosuke Nakamura; Masahide Hikita; Shoko Ogawa; Yasuhiro Takahashi; Takashi Fujishiro
  The FEBS Journal, Volume：287, Number：6, First page：1138, Last page：1154, Mar. 2020, [Reviewed], [Last, Corresponding]
  English, Scientific journal
  DOI：https://doi.org/10.1111/febs.15081
  DOI ID：10.1111/febs.15081, ORCID：63375341
• Identification of IscU residues critical for de novo iron–sulfur cluster assembly　　　　　　　　　　　　　　　
  Naoyuki Tanaka; Eiki Yuda; Takashi Fujishiro; Kei Hirabayashi; Kei Wada; Yasuhiro Takahashi
  Molecular Microbiology, Volume：112, Number：6, First page：1769, Last page：1783, Dec. 2019, [Reviewed]
  Scientific journal
  DOI：https://doi.org/10.1111/mmi.14392
  DOI ID：10.1111/mmi.14392, ISSN：0950-382X, ORCID：62755873, PubMed ID：31532036
• Structure of sirohydrochlorin ferrochelatase SirB: the last of the structures of the class II chelatase family　　　　　　　　　　　　　　　
  Takashi Fujishiro; Yukino Shimada; Ryosuke Nakamura; Miho Ooi
  Dalton Transactions, Volume：48, Number：18, First page：6083, Last page：6090, May 2019, [Reviewed]
  Royal Society of Chemistry ({RSC}), Scientific journal
  DOI：https://doi.org/10.1039/C8DT04727H
  DOI ID：10.1039/C8DT04727H, ISSN：1477-9226, ORCID：54287276, Web of Science ID：WOS:000472449300022
• Distinct roles for U-type proteins in iron–sulfur cluster biosynthesis revealed by genetic analysis of the Bacillus subtilis sufCDSUB operon　　　　　　　　　　　　　　　
  Nao Yokoyama; Chihiro Nonaka; Yukari Ohashi; Masaharu Shioda; Takuya Terahata; Wen Chen; Kotomi Sakamoto; Chihiro Maruyama; Takuya Saito; Eiki Yuda; Naoyuki Tanaka; Takashi Fujishiro; Tomohisa Kuzuyama; Kei Asai; Yasuhiro Takahashi
  Molecular Microbiology, Volume：107, Number：6, First page：688, Last page：703, Mar. 2018, [Reviewed]
  The biosynthesis of iron–sulfur (Fe–S) clusters in Bacillus subtilis is mediated by the SUF-like system composed of the sufCDSUB gene products. This system is unique in that it is a chimeric machinery comprising homologues of E. coli SUF components (SufS, SufB, SufC and SufD) and an ISC component (IscU). B. subtilis SufS cysteine desulfurase transfers persulfide sulfur to SufU (the IscU homologue)
  however, it has remained controversial whether SufU serves as a scaffold for Fe–S cluster assembly, like IscU, or acts as a sulfur shuttle protein, like E. coli SufE. Here we report that reengineering of the isoprenoid biosynthetic pathway in B. subtilis can offset the indispensability of the sufCDSUB operon, allowing the resultant Δsuf mutants to grow without detectable Fe–S proteins. Heterologous bidirectional complementation studies using B. subtilis and E. coli mutants showed that B. subtilis SufSU is interchangeable with E. coli SufSE but not with IscSU. In addition, functional similarity in SufB, SufC and SufD was observed between B. subtilis and E. coli. Our findings thus indicate that B. subtilis SufU is the protein that transfers sulfur from SufS to SufB, and that the SufBCD complex is the site of Fe–S cluster assembly.
  Blackwell Publishing Ltd, English, Scientific journal
  DOI：https://doi.org/10.1111/mmi.13907
  DOI ID：10.1111/mmi.13907, ISSN：1365-2958, SCOPUS ID：85040721939, Web of Science ID：WOS:000426796800003
• Zinc-Ligand Swapping Mediated Complex Formation and Sulfur Transfer between SufS and SufU for Iron-Sulfur Cluster Biogenesis in Bacillus subtilis　　　　　　　　　　　　　　　
  Takashi Fujishiro; Takuya Terahata; Kouhei Kunichika; Nao Yokoyama; Chihiro Maruyama; Kei Asai; Yasuhiro Takahashit
  JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, Volume：139, Number：51, First page：18464, Last page：18467, Dec. 2017, [Reviewed]
  SufU is a zinc-containing protein involved in mobilization of sulfur from SufS for iron-sulfur cluster biogenesis of Bacillus subtilis. Structural basis for the sulfur transfer in SufS-SufU complex was revealed. A zinc-ligand exchange reaction upon SufS-SufU complexation provides a free thiol from Cys41 of SufU as a sulfur acceptor.
  AMER CHEMICAL SOC, English, Scientific journal
  DOI：https://doi.org/10.1021/jacs.7b11307
  DOI ID：10.1021/jacs.7b11307, ISSN：0002-7863, Web of Science ID：WOS:000419082100011
• Mapping the key residues of SufB and SufD essential for biosynthesis of iron-sulfur clusters　　　　　　　　　　　　　　　
  Eiki Yuda; Naoyuki Tanaka; Takashi Fujishiro; Nao Yokoyama; Kei Hirabayashi; Keiichi Fukuyama; Kei Wada; Yasuhiro Takahashi
  SCIENTIFIC REPORTS, Volume：7, Aug. 2017, [Reviewed]
  Biogenesis of iron-sulfur (Fe-S) clusters is an indispensable process in living cells. In Escherichia coli, the SUF biosynthetic system consists of six proteins among which SufB, SufC and SufD form the SufBCD complex, which serves as a scaffold for the assembly of nascent Fe-S cluster. Despite recent progress in biochemical and structural studies, little is known about the specific regions providing the scaffold. Here we present a systematic mutational analysis of SufB and SufD and map their critical residues in two distinct regions. One region is located on the N-terminal side of the beta-helix core domain of SufB, where biochemical studies revealed that Cys254 of SufB (SufB(C254)) is essential for sulfur-transfer from SufE. Another functional region resides at an interface between SufB and SufD, where three residues (SufB(C405), SufB(E434), and SufD(H360)) appear to comprise the site for de novo cluster formation. Furthermore, we demonstrate a plausible tunnel in the beta-helix core domain of SufB through which the sulfur species may be transferred from SufB(C254) to SufB(C405). In contrast, a canonical Fe-S cluster binding motif (CxxCxxxC) of SufB is dispensable. These findings provide new insights into the mechanism of Fe-S cluster assembly by the SufBCD complex.
  NATURE PUBLISHING GROUP, English, Scientific journal
  DOI：https://doi.org/10.1038/s41598-017-09846-2
  DOI ID：10.1038/s41598-017-09846-2, ISSN：2045-2322, Web of Science ID：WOS:000408441600111
• Towards artificial methanogenesis: biosynthesis of the [Fe]-hydrogenase cofactor and characterization of the semisynthetic hydrogenase　　　　　　　　　　　　　　　
  Liping Bai; Takashi Fujishiro; Gangfeng Huang; Urgen Koch; Atsushi Takabayashi; Makio Yokono; Ayumi Tanaka; Tao Xu; Xile Hu; Ulrich Ermler; Seigo Shima
  FARADAY DISCUSSIONS, Volume：198, First page：37, Last page：58, Jun. 2017, [Reviewed]
  The greenhouse gas and energy carrier methane is produced on Earth mainly by methanogenic archaea. In the hydrogenotrophic methanogenic pathway the reduction of one CO2 to one methane molecule requires four molecules of H-2 containing eight electrons. Four of the electrons from two H-2 are supplied for reduction of an electron carrier F-420, which is catalyzed by F-420-reducing [NiFe]-hydrogenase under nickel-sufficient conditions. The same reaction is catalysed under nickel-limiting conditions by [Fe]-hydrogenase coupled with a reaction catalyzed by F-420-dependent methylene tetrahydromethanopterin dehydrogenase. [Fe]-hydrogenase contains an iron-guanylylpyridinol (FeGP) cofactor for H-2 activation at the active site. FeII of FeGP is coordinated to a pyridinol-nitrogen, an acyl-carbon, two CO and a cysteine-thiolate. We report here on comparative genomic analyses of biosynthetic genes of the FeGP cofactor, which are primarily located in a hmd-co-occurring (hcg) gene cluster. One of the gene products is HcgB which transfers the guanosine monophosphate (GMP) moiety from guanosine triphosphate (GTP) to a pyridinol precursor. Crystal structure analysis of HcgB from Methanococcus maripaludis and its complex with 6-carboxymethyl- 3,5-dimethyl-4-hydroxy-2-pyridinol confirmed the physiological guanylyltransferase reaction. Furthermore, we tested the properties of semi-synthetic [Fe]-hydrogenases using the [Fe]-hydrogenase apoenzyme from several methanogenic archaea and a mimic of the FeGP cofactor. On the basis of the enzymatic reactions involved in the methanogenic pathway, we came up with an idea how the methanogenic pathway could be simplified to develop an artificial methanogenesis system.
  ROYAL SOC CHEMISTRY, English, Scientific journal
  DOI：https://doi.org/10.1039/c6fd00209a
  DOI ID：10.1039/c6fd00209a, ISSN：1359-6640, eISSN：1364-5498, Web of Science ID：WOS:000402870300002
• Identification of HcgC as a SAM-Dependent Pyridinol Methyltransferase in [Fe]-Hydrogenase Cofactor Biosynthesis　　　　　　　　　　　　　　　
  Takashi Fujishiro; Liping Bai; Tao Xu; Xiulan Xie; Michael Schick; Joerg Kahnt; Michael Rother; Xile Hu; Ulrich Ermler; Seigo Shima
  ANGEWANDTE CHEMIE-INTERNATIONAL EDITION, Volume：55, Number：33, First page：9648, Last page：9651, Aug. 2016, [Reviewed]
  Previous retrosynthetic and isotope-labeling studies have indicated that biosynthesis of the iron guanylylpyridinol (FeGP) cofactor of [Fe]-hydrogenase requires a methyltransferase. This hypothetical enzyme covalently attaches the methyl group at the 3-position of the pyridinol ring. We describe the identification of HcgC, a gene product of the hcgA-G cluster responsible for FeGP cofactor biosynthesis. It acts as an S-adenosylmethionine (SAM)-dependent methyltransferase, based on the crystal structures of HcgC and the HcgC/SAM and HcgC/S-adenosylhomocysteine (SAH) complexes. The pyridinol substrate, 6-carboxymethyl-5-methyl-4-hydroxy-2-pyridinol, was predicted based on properties of the conserved binding pocket and substrate docking simulations. For verification, the assumed substrate was synthesized and used in a kinetic assay. Mass spectrometry and NMR analysis revealed 6-carboxymethyl-3,5-dimethyl-4-hydroxy-2-pyridinol as the reaction product, which confirmed the function of HcgC.
  WILEY-V C H VERLAG GMBH, English, Scientific journal
  DOI：https://doi.org/10.1002/anie.201604352
  DOI ID：10.1002/anie.201604352, ISSN：1433-7851, eISSN：1521-3773, Web of Science ID：WOS:000383372700033
• A substrate-binding-state mimic of H2O2-dependent cytochrome P450 produced by one-point mutagenesis and peroxygenation of non-native substrates　　　　　　　　　　　　　　　
  Osami Shoji; Takashi Fujishiro; Kousuke Nishio; Yukiko Kano; Hiroshi Kimoto; Shih-Cheng Chien; Hiroki Onoda; Atsushi Muramatsu; Shota Tanaka; Ayumi Hori; Hiroshi Sugimoto; Yoshitsugu Shirob; Yoshihito Watanabe
  CATALYSIS SCIENCE & TECHNOLOGY, Volume：6, Number：15, First page：5806, Last page：5811, 2016, [Reviewed]
  A substrate-binding-state mimic of H2O2-dependent cytochrome P450 that is able to catalyze mono-oxygenation of non-native substrates was constructed by one-point mutagenesis of P450(SP alpha) (CYP152B1). P450(SP alpha), a long-alkyl-chain fatty acid hydroxylase, lacks any general acid-base residue around the heme. The carboxylate group of a fatty acid is thus indispensable for the generation of active species using H2O2. We prepared an A245E mutant to mimic a substrate-binding state by placing a carboxylate group at the active site. The active site structure of the A245E mutant is similar to that of the fatty-acid-bound state of P450(SP alpha) and catalyzes styrene oxidation at a rate of 280 min(-1) (k(cat)), whereas the wild-type enzyme does not show any catalytic activity. More importantly, the same mutation, i.e. the mutation of the highly conserved threonine in P450s to glutamic acid, was also effective in introducing peroxygenase activity into P450BM3, P450(cam), and CYP119. These results indicate that a variety of peroxygenases based on P450s can be constructed by one-point mutagenesis.
  ROYAL SOC CHEMISTRY, English, Scientific journal
  DOI：https://doi.org/10.1039/c6cy00630b
  DOI ID：10.1039/c6cy00630b, ISSN：2044-4753, eISSN：2044-4761, Web of Science ID：WOS:000381431200005
• Reconstitution of [Fe]-hydrogenase using model complexes　　　　　　　　　　　　　　　
  Seigo Shima; Dafa Chen; Tao Xu; Matthew D. Wodrich; Takashi Fujishiro; Katherine M. Schultz; Jorg Kahnt; Kenichi Ataka; Xile Hu
  NATURE CHEMISTRY, Volume：7, Number：12, First page：995, Last page：1002, Dec. 2015, [Reviewed]
  [Fe]-Hydrogenase catalyses the reversible hydrogenation of a methenyltetrahydromethanopterin substrate, which is an intermediate step during the methanogenesis from CO2 and H-2. The active site contains an iron-guanylylpyridinol cofactor, in which Fe2+ is coordinated by two CO ligands, as well as an acyl carbon atom and a pyridinyl nitrogen atom from a 3,4,5,6-substituted 2-pyridinol ligand. However, the mechanism of H-2 activation by [Fe]-hydrogenase is unclear. Here we report the reconstitution of [Fe]-hydrogenase from an apoenzyme using two FeGP cofactor mimics to create semisynthetic enzymes. The small-molecule mimics reproduce the ligand environment of the active site, but are inactive towards H-2 binding and activation on their own. We show that reconstituting the enzyme using a mimic that contains a 2-hydroxypyridine group restores activity, whereas an analogous enzyme with a 2-methoxypyridine complex was essentially inactive. These findings, together with density functional theory computations, support a mechanism in which the 2-hydroxy group is deprotonated before it serves as an internal base for heterolytic H-2 cleavage.
  NATURE PUBLISHING GROUP, English, Scientific journal
  DOI：https://doi.org/10.1038/NCHEM.2382
  DOI ID：10.1038/NCHEM.2382, ISSN：1755-4330, eISSN：1755-4349, Web of Science ID：WOS:000365279200013
• Towards a functional identification of catalytically inactive [Fe]-hydrogenase paralogs　　　　　　　　　　　　　　　
  Takashi Fujishiro; Kenichi Ataka; Ulrich Ermler; Seigo Shima
  FEBS JOURNAL, Volume：282, Number：17, First page：3412, Last page：3423, Sep. 2015, [Reviewed]
  [Fe]-hydrogenase (Hmd), an enzyme of the methanogenic energy metabolism, harbors an iron-guanylylpyridinol (FeGP) cofactor used for H-2 cleavage. The generated hydride is transferred to methenyl-tetrahydromethanopterin (methenyl-H4MPT+). Most hydrogenotrophic methanogens contain the hmd-related genes hmdII and hmdIII. Their function is still elusive. We were able to reconstitute the HmdII holoenzyme of Methanocaldococcus jannaschii with recombinantly produced apoenzyme and the FeGP cofactor, which is a prerequisite for in vitro functional analysis. Infrared spectroscopic and X-ray structural data clearly indicated binding of the FeGP cofactor. Methylene-H4MPT binding was detectable in the significantly altered infrared spectra of the HmdII holoenzyme and in the HmdII apoenzyme-methylene-H4MPT complex structure. The related binding mode of the FeGP cofactor and methenyl-H4MPT+ compared with Hmd and their multiple contacts to the polypeptide highly suggest a biological role in HmdII. However, holo-HmdII did not catalyze the Hmd reaction, not even in a single turnover process, as demonstrated by kinetic measurements. The found inactivity can be rationalized by an increased contact area between the C-and N-terminal folding units in HmdII compared with in Hmd, which impairs the catalytically necessary open-to-close transition, and by an exchange of a crucial histidine to a tyrosine. Mainly based on the presented data, a function of HmdII as Hmd isoenzyme, H2 sensor, FeGP-cofactor storage protein and scaffold protein for FeGP-cofactor biosynthesis could be excluded. Inspired by the recently found binding of HmdII to aminoacyl-tRNA synthetases and tRNA, we tentatively consider HmdII as a regulatory protein for protein synthesis that senses the intracellular methylene-H4MPT concentration.
  WILEY-BLACKWELL, English, Scientific journal
  DOI：https://doi.org/10.1111/febs.13351
  DOI ID：10.1111/febs.13351, ISSN：1742-464X, eISSN：1742-4658, Web of Science ID：WOS:000360629200012
• Protein-pyridinol thioester precursor for biosynthesis of the organometallic acyl-iron ligand in [Fe]-hydrogenase cofactor　　　　　　　　　　　　　　　
  Takashi Fujishiro; Joerg Kahnt; Ulrich Ermler; Seigo Shima
  NATURE COMMUNICATIONS, Volume：6, Apr. 2015, [Reviewed]
  The iron-guanylylpyridinol (FeGP) cofactor of [Fe]-hydrogenase contains a prominent iron centre with an acyl-Fe bond and is the only acyl-organometallic iron compound found in nature. Here, we identify the functions of HcgE and HcgF, involved in the biosynthesis of the FeGP cofactor using structure-to-function strategy. Analysis of the HcgE and HcgF crystal structures with and without bound substrates suggest that HcgE catalyses the adenylylation of the carboxy group of guanylylpyridinol (GP) to afford AMP-GP, and subsequently HcgF catalyses the transesterification of AMP-GP to afford a Cys (HcgF)-S-GP thioester. Both enzymatic reactions are confirmed by in vitro assays. The structural data also offer plausible catalytic mechanisms. This strategy of thioester activation corresponds to that used for ubiquitin activation, a key event in the regulation of multiple cellular processes. It further implicates a nucleophilic attack onto the acyl carbon presumably via an electron-rich Fe(0)-or Fe(I)-carbonyl complex in the Fe-acyl formation.
  NATURE PUBLISHING GROUP, English, Scientific journal
  DOI：https://doi.org/10.1038/ncomms7895
  DOI ID：10.1038/ncomms7895, ISSN：2041-1723, Web of Science ID：WOS:000353704100003
• Structure and function of [Fe]-hydrogenase and biosynthesis of the FeGP cofactor
  Shima, S.; Fujishiro, T.; Ermler, U.
  Biohydrogen, 2015
  Biohydrogen, Scientific journal
  DOI：https://doi.org/10.1515/9783110336733.127
  DOI ID：10.1515/9783110336733.127, ORCID：39616888, SCOPUS ID：84960218575
• A possible iron delivery function of the dinuclear iron center of HcgD in [Fe]-hydrogenase cofactor biosynthesis　　　　　　　　　　　　　　　
  Takashi Fujishiro; Ulrich Ermler; Seigo Shima
  FEBS LETTERS, Volume：588, Number：17, First page：2789, Last page：2793, Aug. 2014, [Reviewed]
  HcgD, a homolog of the ubiquitous Nif3-like protein family, is found in a gene cluster involved in the biosynthesis of the iron-guanylylpyridinol (FeGP) cofactor of [Fe]-hydrogenase. The presented crystal structure and biochemical analyses indicated that HcgD has a dinuclear iron-center, which provides a pronounced binding site for anionic ligands. HcgD contains a stronger and a weaker bound iron; the latter being removable by chelating reagents preferentially in the oxidized state. Therefore, we propose HcgD as an iron chaperone in FeGP cofactor biosynthesis, which might also stimulate investigations on the functionally unknown but physiologically important eukaryotic Nif3-like protein family members.
  Structured summary of protein interactions:
  HcgD and HcgD bind by X-ray crystallography (View interaction) (C) 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
  ELSEVIER SCIENCE BV, English, Scientific journal
  DOI：https://doi.org/10.1016/j.febslet.2014.05.059
  DOI ID：10.1016/j.febslet.2014.05.059, ISSN：0014-5793, eISSN：1873-3468, Web of Science ID：WOS:000340882900005
• Identification of the HcgB Enzyme in [Fe]-Hydrogenase-Cofactor Biosynthesis　　　　　　　　　　　　　　　
  Takashi Fujishiro; Haruka Tamura; Michael Schick; Joerg Kahnt; Xiulan Xie; Ulrich Ermler; Seigo Shima
  ANGEWANDTE CHEMIE-INTERNATIONAL EDITION, Volume：52, Number：48, First page：12555, Last page：12558, Nov. 2013, [Reviewed]
  WILEY-V C H VERLAG GMBH, English, Scientific journal
  DOI：https://doi.org/10.1002/anie.201306745
  DOI ID：10.1002/anie.201306745, ISSN：1433-7851, eISSN：1521-3773, Web of Science ID：WOS:000327582900012
• Crystal Structures of [Fe]-Hydrogenase in Complex with Inhibitory Isocyanides: Implications for the H-2-Activation Site　　　　　　　　　　　　　　　
  Haruka Tamura; Marco Salomone-Stagni; Takashi Fujishiro; Eberhard Warkentin; Wolfram Meyer-Klaucke; Ulrich Ermler; Seigo Shima
  ANGEWANDTE CHEMIE-INTERNATIONAL EDITION, Volume：52, Number：37, First page：9656, Last page：9659, Sep. 2013, [Reviewed]
  WILEY-V C H VERLAG GMBH, English, Scientific journal
  DOI：https://doi.org/10.1002/anie.201305089
  DOI ID：10.1002/anie.201305089, ISSN：1433-7851, eISSN：1521-3773, Web of Science ID：WOS:000323829600011
• Crystal structures of [fe]-hydrogenase in complex with inhibitory isocyanides: implications for the h2 -activation site.　　　　　　　　　　　　　　　
  Tamura, Haruka; Salomone-Stagni, Marco; Fujishiro, Takashi; Warkentin, Eberhard; Meyer-Klaucke, Wolfram; Ermler, Ulrich; Shima, Seigo
  Angewandte Chemie (International ed. in English), Volume：52, Number：37, 2013
  Scientific journal
  DOI：https://doi.org/10.1002/anie.201305089
  DOI ID：10.1002/anie.201305089, ISSN：1521-3773, ORCID：10195685
• Chiral-Substrate-Assisted Stereoselective Epoxidation Catalyzed by H2O2-Dependent Cytochrome P450SPa　　　　　　　　　　　　　　　
  Takashi Fujishiro; Osami Shoji; Norifumi Kawakami; Takahiro Watanabe; Hiroshi Sugimoto; Yoshitsugu Shiro; Yoshihito Watanabe
  CHEMISTRY-AN ASIAN JOURNAL, Volume：7, Number：10, First page：2286, Last page：2293, Oct. 2012, [Reviewed]
  The stereoselective epoxidation of styrene was catalyzed by H2O2-dependent cytochrome P450SPa in the presence of carboxylic acids as decoy molecules. The stereoselectivity of styrene oxide could be altered by the nature of the decoy molecules. In particular, the chirality at the a-positions of the decoy molecules induced a clear difference in the chirality of the product: (R)-ibuprofen enhanced the formation of (S)-styrene oxide, whereas (S)-ibuprofen preferentially afforded (R)-styrene oxide. The crystal structure of an (R)-ibuprofen-bound cytochrome P450SPa (resolution 1.9 angstrom) revealed that the carboxylate group of (R)-ibuprofen served as an acidbase catalyst to initiate the epoxidation. A docking simulation of the binding of styrene in the active site of the (R)-ibuprofen-bound form suggested that the orientation of the vinyl group of styrene in the active site agreed with the formation of (S)-styrene oxide.
  WILEY-V C H VERLAG GMBH, English, Scientific journal
  DOI：https://doi.org/10.1002/asia.201200250
  DOI ID：10.1002/asia.201200250, ISSN：1861-4728, Web of Science ID：WOS:000308877400018
• Construction of biocatalysts using the myoglobin scaffold for the synthesis of indigo from indole　　　　　　　　　　　　　　　
  Jiakun Xu; Osami Shoji; Takashi Fujishiro; Takahiro Ohki; Takafumi Ueno; Yoshihito Watanabe
  CATALYSIS SCIENCE & TECHNOLOGY, Volume：2, Number：4, First page：739, Last page：744, 2012, [Reviewed]
  Hydrogen peroxide-dependent oxidation of indole producing indigo has been catalyzed by engineered myoglobins (Mbs). We have investigated a series of H64D mutants of Mbs in which Asp-64 accelerates the reaction of the Mbs with hydrogen peroxide and stabilizes the resulting active species: compound I (iron oxo-ferryl porphyrin pi-cation radical). The catalytic activity of mutants depends on the amino acids at 68 and 107 positions and the H64D/V68I/I107A mutant gave the highest catalytic activity, suggesting that the side chains of Ile-68 and Ala-107 provide a hydrophobic environment suitable for indole binding. The k(cat) of the H64D/V68I/I107A mutant achieved 72 min(-1), indicating that even Mb can be transformed into a biocatalyst.
  ROYAL SOC CHEMISTRY, English, Scientific journal
  DOI：https://doi.org/10.1039/c2cy00427e
  DOI ID：10.1039/c2cy00427e, ISSN：2044-4753, Web of Science ID：WOS:000302062900011
• Crystal Structure of H2O2-dependent Cytochrome P450(SP alpha) with Its Bound Fatty Acid Substrate INSIGHT INTO THE REGIOSELECTIVE HYDROXYLATION OF FATTY ACIDS AT THE alpha POSITION　　　　　　　　　　　　　　　
  Takashi Fujishiro; Osami Shoji; Shingo Nagano; Hiroshi Sugimoto; Yoshitsugu Shiro; Yoshihito Watanabe
  JOURNAL OF BIOLOGICAL CHEMISTRY, Volume：286, Number：34, First page：29941, Last page：29950, Aug. 2011, [Reviewed]
  Cytochrome P450(SP alpha) (CYP152B1) isolated from Sphingomonas paucimobilis is the first P450 to be classified as a H2O2-dependent P450. P450(SP alpha) hydroxylates fatty acids with high alpha-regioselectivity. Herein we report the crystal structure of P450(SP alpha) with palmitic acid as a substrate at a resolution of 1.65 angstrom. The structure revealed that the C-alpha of the bound palmitic acid in one of the alternative conformations is 4.5 angstrom from the heme iron. This conformation explains the highly selective alpha-hydroxylation of fatty acid observed in P450(SP alpha). Mutations at the active site and the F-G loop of P450(SP alpha) did not impair its regioselectivity. The crystal structures of mutants (L78F and F288G) revealed that the location of the bound palmitic acid was essentially the same as that in the WT, although amino acids at the active site were replaced with the corresponding amino acids of cytochrome P450(BS alpha) (CYP152A1), which shows beta-regioselectivity. This implies that the high regioselectivity of P450(SP alpha) is caused by the orientation of the hydrophobic channel, which is more perpendicular to the heme plane than that of P450(BS alpha)
  AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, English, Scientific journal
  DOI：https://doi.org/10.1074/jbc.M111.245225
  DOI ID：10.1074/jbc.M111.245225, ISSN：0021-9258, Web of Science ID：WOS:000294046600053
• Non-covalent modification of the active site of cytochrome P450 for inverting the stereoselectivity of monooxygenation　　　　　　　　　　　　　　　
  Takashi Fujishiro; Osami Shoji; Yoshihito Watanabe
  TETRAHEDRON LETTERS, Volume：52, Number：3, First page：395, Last page：397, Jan. 2011, [Reviewed]
  The enantioselectivity in the sulfoxidation of thioanisole catalyzed by cytochrome P450(BS beta) with a decoy molecule, a dummy molecule of the natural substrate, can be inverted by changing the structure of the decoy molecule. The methodology demonstrated herein shows the potential for controlling the stereoselectivity of biocatalysts without any mutagenesis. (C) 2010 Elsevier Ltd. All rights reserved.
  PERGAMON-ELSEVIER SCIENCE LTD, English, Scientific journal
  DOI：https://doi.org/10.1016/j.tetlet.2010.11.048
  DOI ID：10.1016/j.tetlet.2010.11.048, ISSN：0040-4039, Web of Science ID：WOS:000286862200009
• Understanding substrate misrecognition of hydrogen peroxide dependent cytochrome P450 from Bacillus subtilis　　　　　　　　　　　　　　　
  Osami Shoji; Takashi Fujishiro; Shingo Nagano; Shota Tanaka; Takuya Hirose; Yoshitsugu Shiro; Yoshihito Watanabe
  JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY, Volume：15, Number：8, First page：1331, Last page：1339, Nov. 2010, [Reviewed]
  Cytochrome P450(Bs beta) a H(2)O(2)-dependent cytochrome P450 catalyzing the hydroxylation of long-alkyl-chain fatty acids, lacks the general acid base residue around the heme, which is indispensable for the efficient generation of the active species using H(2)O(2) On the basis of the crystal structure of the palmitic acid bound form of cytochrome P450(Bs beta), it was suggested that the role of the general acid base function was provided by the carboxylate group of fatty acids The participation of the carboxylate group of the substrate was supported by the fact that cytochrome P450(Bs beta) can catalyze oxidations of nonnatural substrates such as styrene and ethylbenzene in the presence of a series of short-alkyl-chain carboxylic acids as a dummy molecule of fatty acid We refer to a series of short-alkyl-chain carboxylic acids as a "decoy molecule" As shown here, we have clarified the crystal structure of the decoy-molecule-bound form and elucidated that the location of its carboxylate group is virtually the same as that of palmitic acid in the heme cavity, indicating that the carboxylate group of the decoy molecule serves as the general acid base catalyst This result further confirms that the role of the acid base function is satisfied by the carboxylate group of the substrates In addition, the structure analysis of the substrate-free form has clarified that no remarkable structural change is induced by the binding of the decoy molecule as well as fatty acid Consequently, whether the carboxylate group is positioned in the active site provides the switching mechanism of the catalytic cycle of cytochrome P450(Bs beta)
  SPRINGER, English, Scientific journal
  DOI：https://doi.org/10.1007/s00775-010-0692-4
  DOI ID：10.1007/s00775-010-0692-4, ISSN：0949-8257, Web of Science ID：WOS:000285264900016
• Aromatic C-H bond hydroxylation by P450 peroxygenases: a facile colorimetric assay for monooxygenation activities of enzymes based on Russig's blue formation (vol 15, pg 1109, 2010)　　　　　　　　　　　　　　　
  Osami Shoji; Christian Wiese; Takashi Fujishiro; Chikako Shirataki; Bernhard Wuensch; Yoshihito Watanabe
  JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY, Volume：15, Number：7, First page：1171, Last page：1171, Sep. 2010, [Reviewed]
  SPRINGER, English
  DOI：https://doi.org/10.1007/s00775-010-0679-1
  DOI ID：10.1007/s00775-010-0679-1, ISSN：0949-8257, Web of Science ID：WOS:000281597900018
• Aromatic C-H bond hydroxylation by P450 peroxygenases: a facile colorimetric assay for monooxygenation activities of enzymes based on Russig's blue formation　　　　　　　　　　　　　　　
  Osami Shoji; Christian Wiese; Takashi Fujishiro; Chikako Shirataki; Bernhard Wuensch; Yoshihito Watanabe
  JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY, Volume：15, Number：7, First page：1109, Last page：1115, Sep. 2010, [Reviewed]
  Aromatic C-H bond hydroxylation of 1-methoxynaphthalene was efficiently catalyzed by the substrate misrecognition system of the hydrogen peroxide dependent cytochrome P450(BS beta) (CYP152A1), which usually catalyzes hydroxylation of long-alkyl-chain fatty acids. Very importantly, the hydroxylation of 1-methoxynaphthalene can be monitored by a color change since the formation of 4-methoxy-1-naphthol was immediately followed by its further oxidation to yield Russig's blue. Russig's blue formation allows us to estimate the peroxygenation activity of enzymes without the use of high performance liquid chromatography, gas chromatography, and nuclear magnetic resonance measurements.
  SPRINGER, English, Scientific journal
  DOI：https://doi.org/10.1007/s00775-010-0671-9
  DOI ID：10.1007/s00775-010-0671-9, ISSN：0949-8257, Web of Science ID：WOS:000281597900012
• Hydrogen peroxide dependent monooxygenations by tricking the substrate recognition of cytochrome P450(BS beta)　　　　　　　　　　　　　　　
  Osami Shoji; Takashi Fujishiro; Hiroshi Nakajima; Misa Kim; Shingo Nagano; Yoshitsugu Shiro; Yoshihito Watanabe
  ANGEWANDTE CHEMIE-INTERNATIONAL EDITION, Volume：46, Number：20, First page：3656, Last page：3659, 2007, [Reviewed]
  WILEY-V C H VERLAG GMBH, English, Scientific journal
  DOI：https://doi.org/10.1002/anie.200700068
  DOI ID：10.1002/anie.200700068, ISSN：1433-7851, Web of Science ID：WOS:000246651600008
• Control of the catalytic activity of cytochrome P450BSβ by various guest molecules with carboxyl group　　　　　　　　　　　　　　　
  Shoji, O.; Fujishiro, T.; Nakajima, H.; Watanabe, Y.
  Polymer Preprints, Japan, Volume：55, Number：1, 2006
  Scientific journal
  ORCID：9066370, SCOPUS ID：33749855729

• The minimal SUF system can substitute for the canonical iron-sulfur cluster biosynthesis systems by using inorganic sulfide as the sulfur source　　　　　　　　　　　　　　　
  Maya Murata; Taichi Murakami; Eiki Yuda; Nanami Mukai; Xintong Zheng; Natsumi Kurachi; Sachiko Mori; Shoko Ogawa; Kouhei Kunichika; Takashi Fujishiro; Kei Wada; Yasuhiro Takahashi
  21 Mar. 2024
  DOI：https://doi.org/10.1101/2024.03.20.586028
  DOI ID：10.1101/2024.03.20.586028, ORCID put code：156013284
• Sulfur-mobilizing Enzymes Involved in Iron-sulfur Cluster Biosynthesis: Shared Structural Features and Functional Diversity　　　　　　　　　　　　　　　
  FUJISHIRO Takashi; NAKAMURA Ryosuke; TAKAHASHI Yasuhiro
  Seibutsu Butsuri, Volume：61, Number：3, First page：180, Last page：182, 2021
  The Biophysical Society of Japan General Incorporated Association, Japanese
  DOI：https://doi.org/10.2142/biophys.61.180
  DOI ID：10.2142/biophys.61.180, ISSN：0582-4052, CiNii Articles ID：130008045148
• 埼玉新聞サイ・テクこらむ–金属と生命、そして進化　　　　　　　　　　　　　　　
  藤城貴史
  09 Oct. 2019, [Invited], [Lead, Corresponding]
• Chem-Station第１３３回スポットライトリサーチ「システインから無機硫黄を取り出す酵素反応の瞬間を捉える」　　　　　　　　　　　　　　　
  藤城 貴史
  Jan. 2018, [Invited]
  Japanese, Introduction commerce magazine
• Bioinorganic chemistry on biosynthesis of metal clusters in metalloproteins　　　　　　　　　　　　　　　
  藤城 貴史
  Number：9, First page：17, Last page：20, 2018
  Metals are essential for all kinds of living organisms and utilized by metalloproteins as their metal cofactors. Because of their unique properties, metalloproteins and their metal cofactors have been extensively studied by using multidisciplinary approaches in "bioinorganic chemistry". This review briefly highlights how biosynthesis of biological metal clusters in metalloproteins can be investigated by taking an example of our recent results in studying biosynthesis of the iron-sulfur cluster.
  Comprehensive Analysis Center for Science, Saitama University, Japanese
  DOI：https://doi.org/10.24561/00018467
  DOI ID：10.24561/00018467, CiNii Articles ID：120006577230, CiNii Books ID：AA12504266
• 留学体験記「マックスプランク陸生微生物学研究所留学体験記〜化学から生物学へ〜」　　　　　　　　　　　　　　　
  藤城 貴史
  Volume：55, Oct. 2017, [Invited]
  Japanese, Others
• [Fe]-ヒドロゲナーゼの活性中心コファクターと立体構造解析に基づくその生合成酵素の同定　　　　　　　　　　　　　　　
  嶋 盛吾; 藤城 貴史
  Volume：76, First page：25, Last page：29, Oct. 2016, [Reviewed], [Invited]
  Japanese, Introduction scientific journal
• 埼玉新聞サイ・テクこらむ–タンパク質の構造と役割　　　　　　　　　　　　　　　
  藤城 貴史
  Feb. 2016, [Invited]
  Japanese, Introduction commerce magazine
• 酵素立体構造からの機能解読　　　　　　　　　　　　　　　
  藤城 貴史; 嶋 盛吾
  Volume：92, First page：676, Last page：676, Dec. 2014, [Invited]
  Japanese, Introduction scientific journal
• Identification of the Hcg enzymes in biosynthesis of [Fe]-hydrogenase cofactor by a structural genomics-based approach　　　　　　　　　　　　　　　
  Takashi Fujishiro; Haruka Tamura; Michael Schick; Joerg Kahnt; Xiulan Xie; Ulrich Ermler; Seigo Shima
  JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY, Volume：19, First page：S767, Last page：S767, Aug. 2014
  SPRINGER, English, Summary international conference
  ISSN：0949-8257, eISSN：1432-1327, Web of Science ID：WOS:000339874700152
• Biochemical and X-ray crystal structure analyses of Hcg proteins involved in biosynthesis of the FeGP cofactor of [Fe]-hydrogenase　　　　　　　　　　　　　　　
  T. Fujishiro; H. Tamura; M. Schick; J. Kahnt; S. Shima; X. Xie; U. Ermler; S. Shima
  JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY, Volume：19, First page：S223, Last page：S223, Mar. 2014
  SPRINGER, English, Summary international conference
  ISSN：0949-8257, eISSN：1432-1327, Web of Science ID：WOS:000332835300164
• Alteration of substrate specificity of heme enzymes using decoy molecules
  Fujishiro Takashi
  Bull. Jpn. Soc. Coord. Chem., Volume：51, Number：51, First page：82, Last page：84, 2008, [Invited]
  Japanese, Introduction scientific journal
  ISSN：1882-6954, CiNii Articles ID：40016172578

• Bioorganometallic Chemistry　　　　　　　　　　　　　　　
  Takashi Fujishiro (Edited by Wolfgang Weigand; Ulf-Peter Apfel), [Joint work], Part I chapter 2 "Hydrogen development" [Fe]-hydrogenase
  De Gruyter, May 2020
  共同研究・競争的資金等ID：11131676
• Biohydrogen, (Edited by Prof. Dr. Matthias Rögner)　　　　　　　　　　　　　　　
  Seigo Shima; Takashi Fujishiro; Ulrich Ermler, [Contributor], Chapter 6 "Structure and function of [Fe]-hydrogenase and biosynthesis of the FeGP cofactor"
  De Gruyter, 2015
• Construction of monooxygenation systems using hydrogen peroxide-dependent cytochrome P450s　　　　　　　　　　　　　　　
  Takashi Fujishiro, [Single work]
  2011
  English
  CiNii Books：http://ci.nii.ac.jp/ncid/BB12440635
  CiNii Books ID：BB12440635

• AI 技術で紐解く CO 触媒酵素のガストンネルの変遷と炭素固定進化　　　　　　　　　　　　　　　
  大前公保; 藤城貴史; 石井俊一; 鈴木志野
  Mar. 2025
  Mar. 2025 - Mar. 2025, Japanese, Oral presentation
  共同研究・競争的資金等ID：42101092
• 金属-テトラピロール補因子の生合成酵素キラターゼの構造と機能の進化の追跡　　　　　　　　　　　　　　　
  小川翔子; 小山田勇真; 藤城貴史
  Nov. 2024
  Nov. 2024 - Nov. 2024, Japanese, Poster presentation
  共同研究・競争的資金等ID：25988841
• Challenges for molecular design of Fe-S enzymes functioning as an artificial Ni,Fe-CO dehydrogenase　　　　　　　　　　　　　　　
  Takashi Fujishiro
  The 2nd CO world Plenary Meeting, Aug. 2024
  Jul. 2024 - Aug. 2024, English, Oral presentation
  共同研究・競争的資金等ID：42101092
• 三重結合アミノ酸生合成系を改変したクリックケミストリー指向型ペプチド合成系の構築　　　　　　　　　　　　　　　
  藤城貴史
  Jul. 2024
  Jul. 2024 - Jul. 2024, Japanese, Oral presentation
  共同研究・競争的資金等ID：42101093
• PLP依存型システイン脱硫酵素IscSの活性の起源の解明　　　　　　　　　　　　　　　
  吉田竜河; 國近航平; 佐々木大和; 林菜海; 山川誠; 岩永朋子; 和田啓; 高橋康弘; 藤城貴史
  Jun. 2024
  Jun. 2024 - Jun. 2024, Japanese, Poster presentation
• Study on E. coli recombinant metalloproteins with use of synthetic genes　　　　　　　　　　　　　　　
  Takashi Fujishiro
  2024 annual meeting of Japan Scociety for Bioscience, Biotechnology, and Agrochemistry, Mar. 2024, [Invited]
  Mar. 2024 - Mar. 2024, Invited oral presentation
  共同研究・競争的資金等ID：42101092
• Insights into structural evolution of Ni,Fe-CO dehydrogenases and hybrid cluster proteins　　　　　　　　　　　　　　　
  Takashi Fujishiro
  Joint CO world & 12th ELSI Symposium “EMERGENCE AND DETECTION OF LIFE, Jan. 2024, [Invited]
  Jan. 2024 - Jan. 2024, English, Invited oral presentation
  共同研究・競争的資金等ID：42101092
• Understanding of substrate recognition mechanism of nickelochelatase CfbA and modification of its tetrapyrrole substrate selectivity　　　　　　　　　　　　　　　
  Shoko Ogawa; Masahide Hikita; Yuma Oyamada; Takashi Fujishiro
  Sep. 2023
  Sep. 2023 - Sep. 2023, English, Poster presentation
• キラターゼHemHを鋳型として用いた金属センサー人工蛍光タンパク質の創製　　　　　　　　　　　　　　　
  宍戸萌恵; 藤城貴史
  Sep. 2023
  Sep. 2023 - Sep. 2023, Japanese, Poster presentation
• ニッケルキラターゼCfbAの基質選択機構の解明　　　　　　　　　　　　　　　
  小川翔子; 小山田勇真; 引田理英; 藤城貴史
  Sep. 2023
  Sep. 2023 - Sep. 2023, Japanese, Poster presentation
• Understanding and rational alteration of tetrapyrrole substrate selectivity of nickelochelatase CfbA　　　　　　　　　　　　　　　
  Shoko Ogawa; Masahide Hikita; Yuuma Oyamada; Takashi Fujishiro
  ICBIC20, Jul. 2023
  Jul. 2023 - Jul. 2023, English, Poster presentation
• 疎水空間と金属結合部位を併せ持つHemHを鋳型とした金属センサー型人工蛍光タンパク質の合成　　　　　　　　　　　　　　　
  宍戸萌恵; 小山田勇真; 藤城貴史
  May 2023
  May 2023 - May 2023, Japanese, Poster presentation
• Structural Analysis of Class II Hybrid Cluster Protein (HCP) from Escherichia coli　　　　　　　　　　　　　　　
  Takashi Fujishiro
  10th Asian Biological Inorganic Inorganic Chemistry Conference (AsBIC10), Dec. 2022, [Invited]
  Nov. 2022 - Dec. 2022, English, Invited oral presentation
  共同研究・競争的資金等ID：37071571
• Structural basis for biosynthesis of a [4Fe-4S] cluster from two [2Fe-2S] clusters on the IscU enzyme in ISC machinery　　　　　　　　　　　　　　　
  Kouhei Kunichika; Ryosuke Nakamura; Takashi Fujishiro; Yasuhiro Takahashi
  10th Asian Biological Inorganic Inorganic Chemistry Conference (AsBIC10), Nov. 2022
  Nov. 2022 - Dec. 2022, English, Poster presentation
  共同研究・競争的資金等ID：37071572
• Structures of Nickelochelatase CfbA with Non-Physiological Porphyrinoids　　　　　　　　　　　　　　　
  Shoko Ogawa; Masahide Hikita; Yuuma Oyamada; Takashi Fujishiro
  10th Asian Biological Inorganic Inorganic Chemistry Conference (AsBIC10), Nov. 2022
  Nov. 2022 - Dec. 2022, English, Poster presentation
  共同研究・競争的資金等ID：25988841
• 基質選択性の改変に向けた鉄キラターゼSirBの活性部位の分子設計　　　　　　　　　　　　　　　
  藤城貴史; 小山田勇真; 小川翔子
  Nov. 2022
  Nov. 2022 - Nov. 2022, Japanese, Poster presentation
  共同研究・競争的資金等ID：25988841
• ２つの異なるタイプのシステインデスルフラーゼと基質類似分子との反応　　　　　　　　　　　　　　　
  大塚穂乃; 中村亮裕; 小川翔子; 藤城貴史; 高橋康弘
  Nov. 2022
  Nov. 2022 - Nov. 2022, Japanese, Poster presentation
  共同研究・競争的資金等ID：37071572
• メタン生成古細菌のSufB2C2複合体は鉄硫黄クラスター生合成系SUFマシナリーのプロトタイプである　　　　　　　　　　　　　　　
  村上太一; 村田真耶; 藤城貴史; 高橋康弘
  Nov. 2022
  Nov. 2022 - Nov. 2022, Japanese, Poster presentation
  共同研究・競争的資金等ID：37071572
• 鉄硫黄クラスター生合成に関わるSUF、SUF-like、ISC系の 硫黄供給システムの酸化ストレス耐性の比較　　　　　　　　　　　　　　　
  槇千智; 室賀 直来; 寺畑 拓也; 島田 侑希乃; 國近 航平; 中村 亮裕; 藤城 貴史; 高橋 康弘
  Nov. 2022
  Nov. 2022 - Nov. 2022, Japanese, Poster presentation
  共同研究・競争的資金等ID：37071572
• Actions of two distinct types of PLP-dependent cysteine desulfurase enzymes with substrate L-cysteine and inhibitors　　　　　　　　　　　　　　　
  Takashi Fujishiro
  Sep. 2022, [Invited]
  Sep. 2022 - Sep. 2022, English, Invited oral presentation
  共同研究・競争的資金等ID：12335178
• Structural insights into porphyrinoid substrate selectivity of chelatase CfbA　　　　　　　　　　　　　　　
  Shoko Ogawa; Yuma Oyamada; Masahide Hikita; Takashi Fujishiro
  Sep. 2022
  Sep. 2022 - Sep. 2022, English, Poster presentation
  共同研究・競争的資金等ID：25988841
• Structural insights into biosynthesis of a [4Fe-4S] cluster via coupling of two adjacent [2Fe−2S] clusters in the IscU enzyme　　　　　　　　　　　　　　　
  Kouhei Kunichika; Ryosuke Nakamura; Takashi Fujishiro; Yasuhiro Takahashi
  Sep. 2022
  Sep. 2022 - Sep. 2022, English, Poster presentation
  共同研究・競争的資金等ID：12335178
• 祖先型キラターゼCfbAの基質選択性の構造に基づく理解　　　　　　　　　　　　　　　
  小川翔子; 小山田勇真; 引田理英; 藤城貴史
  Jun. 2022
  Jun. 2022 - Jun. 2022, Japanese, Oral presentation
  共同研究・競争的資金等ID：25988841
• 鉄硫黄クラスター：鉄と硫黄からなる無機集合体の生合成と多様性　　　　　　　　　　　　　　　
  藤城貴史
  Mar. 2022, [Invited]
  Mar. 2022 - Mar. 2022, Japanese, Invited oral presentation
• Sulfur acquisition mechanism for iron-sulfur cluster biosynthesis　　　　　　　　　　　　　　　
  Takashi Fujishiro; Yasuhiro Takahashi
  Pacifichem2021, Dec. 2021, [Invited]
  Dec. 2021 - Dec. 2021, English, Invited oral presentation
• 金属ポルフィリン型補因子の生合成鍵酵素キラターゼの分子進化　　　　　　　　　　　　　　　
  藤城貴史
  Nov. 2021, [Invited]
  Nov. 2021 - Nov. 2021, Japanese, Invited oral presentation
• 鉄キラターゼSirBのニッケルキラターゼへの機能改変　　　　　　　　　　　　　　　
  小山田 勇真; 小川 翔子; 藤城 貴史
  Nov. 2021
  Nov. 2021 - Nov. 2021, Japanese, Poster presentation
• 鉄硫黄クラスター生合成の硫黄輸送に関わる分子種の比較　　　　　　　　　　　　　　　
  槇 千智; 室賀 直来; 寺畑 拓也; 島田 侑希乃; 國近 航平; 中村 亮裕; 藤城 貴史; 高橋 康弘
  Nov. 2021
  Nov. 2021 - Nov. 2021, Japanese, Poster presentation
• 鉄硫黄クラスター生合成に関与するSufEおよびSufUの硫黄輸送状態の比較　　　　　　　　　　　　　　　
  槇 千智; 室賀 直来; 寺畑 拓也; 島田 侑希乃; 國近 航平; 中村 亮裕; 藤城 貴史; 高橋 康弘
  Nov. 2021
  Nov. 2021 - Nov. 2021, Japanese, Poster presentation
• ニッケルキラターゼCfbAの基質選択性の理解　　　　　　　　　　　　　　　
  小川 翔子; 小山田 勇真; 引田 理英; 藤城 貴史
  Nov. 2021
  Nov. 2021 - Nov. 2021, Japanese, Poster presentation
• 補酵素F430の生合成系のニッケルキラターゼの触媒機構の解明　　　　　　　　　　　　　　　
  藤城 貴史; 小川 翔子
  Nov. 2021
  Nov. 2021 - Nov. 2021, Japanese, Poster presentation
• Structure and properties of Escherichia coli Hybrid cluster protein (HCP) harboring an Fe-S-O cluster　　　　　　　　　　　　　　　
  Takashi Fujishiro; Miho Ooi; Kyosei Takaoka
  Sep. 2021
  Sep. 2021 - Sep. 2021, English, Oral presentation
• 鉄硫黄クラスター生合成足場タンパク質IscUにおける[2Fe-2S]から[4Fe-4S]へのクラスター変換の構造基盤　　　　　　　　　　　　　　　
  國近 航平; 中村 亮裕; 藤城 貴史; 高橋 康弘
  Jun. 2021
  Jun. 2021 - Jun. 2021
• PLP依存型酵素システインデスルフラーゼSufSに対する抗生物質シクロセリンの鏡像異性体間で異なる阻害反応　　　　　　　　　　　　　　　
  中村 亮裕; 小川 翔子; 小松 茉里佳; 藤城 貴史; 高橋 康弘
  Jun. 2021
  Jun. 2021 - Jun. 2021
• 鉄硫黄クラスター生合成マシナリーのシステイン脱硫酵素のシクロセリンによる阻害機構の解明　　　　　　　　　　　　　　　
  小川 翔子; 中村 亮裕; 小松 茉里佳; 引田 理英; 藤城 貴史; 高橋 康弘
  Sep. 2020
  Sep. 2020 - Sep. 2020
• 鉄硫黄クラスター生合成足場タンパク質IscUの[2Fe-2S]クラスター集積型二量体構造とその意義　　　　　　　　　　　　　　　
  國近 航平; 藤城 貴史; 高橋 康弘
  Sep. 2020
  Sep. 2020 - Sep. 2020
• Characterization of Fe-S-O hybrid cluster protein found in facultative anaerobes　　　　　　　　　　　　　　　
  Takashi Fujishiro, Miho Ooi, Yasuhiro Takahashi
  The 100th Chemical Soceity of Japan Annual Meeting, Mar. 2020
  Mar. 2020 - Mar. 2020, Japanese, Oral presentation
• Elucidation of inhibition mechanism of PLP-dependent SufS enzyme in an Fe-S cluster biosynthetic machinery by cycloserine　　　　　　　　　　　　　　　
  Shoko Ogawa, Ryosuke Nakamura, Marika Komatsu, Masahide Hikita, Takashi Fujishiro, Yasuhiro Takahashi
  The 100th Chemical Society of Japan Annual Meeting, Mar. 2020
  Mar. 2020 - Mar. 2020, Japanese, Oral presentation
• A biologically unusual [4Fe-4S] cluster coordination found in a cysteine-degradating enzyme　　　　　　　　　　　　　　　
  Takashi Fujishiro; Daichi Yamanouchi; Yuuki Kamioka; Yasuhiro Takahashi
  The 69th Annual Meeting of the Coordination Chemistry of Japan, Sep. 2019, [Domestic conference]
  English, Oral presentation
• A role of zinc-protein SufU in Fe-S cluster biosynthetic SUF-like machinery　　　　　　　　　　　　　　　
  Takashi Fujishiro; Takuya Terahata; Yukino Shimada; Yasuhiro Takahashi
  The 92nd annual meeting of the Japanese biochemical society, Sep. 2019, [Domestic conference]
  Japanese, Poster presentation
• Structure-function relationship of sirohydrochlorin ferrochelatase SirB　　　　　　　　　　　　　　　
  Takashi Fujishiro; Yukino Shimada; Ryosuke Nakamura; Miho Ooi
  19th International Conference on Biological Inorganic Chemistry (ICBIC19), Aug. 2019, [International conference]
  English, Poster presentation
• X-ray crystallographic snapshots of sulfur mobilization to iron-sulfur cluster biosynthesis　　　　　　　　　　　　　　　
  Fujishiro Takashi
  The International Symposium on Bioinorganic Chemistry 2018, Nov. 2018, [Invited], [International conference]
  English, Invited oral presentation
• Exploring an unusual Cys-rich motif-containing Fe-S enzyme　　　　　　　　　　　　　　　
  T. Fujishiro; Mio Ohi; Yasuhiro Takahashi
  The 91st annual meeting of teh Japanese Biochemical Society, Sep. 2018, [Domestic conference]
  Japanese, Poster presentation
• [Fe]-Hydrogenase-cofactor biosynthesis　　　　　　　　　　　　　　　
  T. Fujishiro
  ICCC2018, Jul. 2018, [Invited], [International conference]
  English, Invited oral presentation
• Zinc-mediated sulfur acquisition by SufS-SufU complex involved in biosynthesis of iron-sulfur cluster in Bacillus subtilis　　　　　　　　　　　　　　　
  T. Fujishiro; T. Terahata; K. Kunichika; N. Yokoyama; C. Maruyama; K. Asai; Y. Takahashi
  The 68th conference of Japan Society of Coordination Chemistry, Jul. 2018, [Domestic conference]
  English, Oral presentation
• X-ray crystallographic snapshot of SufS-SufU complex in iron-sulfur cluster biosynthetic SUF-like machinery　　　　　　　　　　　　　　　
  T. Fujishiro; T. Terahata; K. Kunichika; N. Yokoyama; C. Maruyama; K. Asai; Y. Takahashi
  The 18th annual meeting of the Protein Science Society of Japan, Jun. 2018, [Domestic conference]
  Japanese, Poster presentation
• X-ray crytallographic analysis of sulfur-transferring complex in iron-sulfur cluster biosynthesis of Bacillus subtilis　　　　　　　　　　　　　　　
  T. Fujishiro; T. Terahata; K. Kunichika; N. Yokoyama; C. Maruyama; K. Asai; Y. Takahashi
  The 98th annual meeting of the Chemical Society of Japan, Mar. 2018, [Domestic conference]
  English, Oral presentation
• Structure-guided functional analysis of metallochelatase-like protein from Bacillus subtilis　　　　　　　　　　　　　　　
  T. Fujishiro; K. Asai; Y. Takahashi
  ConBio2017, Dec. 2017, [Domestic conference]
  Japanese, Poster presentation
• The NIF machinery for iron-sulfur cluster biogenesis　　　　　　　　　　　　　　　
  Takashi Fujishiro; Yasuhiro Takahashi
  The 17th annual meeting of the Protein Science Society of Japan, Jun. 2017, [Invited], [Domestic conference]
  English, Invited oral presentation
• Protein-protein interaction analysis of iron-sulfur cluster biosynthetic protein IscA and ferredoxin FdxN　　　　　　　　　　　　　　　
  T. Fujishiro; C. Ohno; Y. Kamioka; Y. Takahashi
  The 97th annual meeting of the Chemical Society of Japan, Mar. 2017, [Domestic conference]
  Japanese, Oral presentation
• The iron-sulfur cluster biosynthesis　　　　　　　　　　　　　　　
  T. Fujishiro; Y. Takahashi
  ISSP-Workshop, Mar. 2017, [Invited], [Domestic conference]
  English, Invited oral presentation
• Elucidation of structure-function relationshipof cysteine desulfurase NifS in NIF machinery for iron-sulfur cluster biosynthesis by co-crystallization and ligand screening　　　　　　　　　　　　　　　
  T. Fujishiro; R. Nakamura; S. Sato; Y. Takahashi
  The 43rd symposium on biomolecular science, Jun. 2016, [Domestic conference]
  Japanese, Oral presentation
• Towards a creation of a whole cell catalyst for H2 production by heterologous expression of [FeFe]-hydrogenase and its related genes in Escherichia coli　　　　　　　　　　　　　　　
  Takashi Fujishiro; Yasuhiro Takahashi
  The 96th annual meeting of the Chemical Society of Japan, Mar. 2016, [Domestic conference]
  Japanese, Poster presentation
• Identification of Hcg enzymes in volved in [Fe]-hydrogenase cofactor biosynthesis by using structural information　　　　　　　　　　　　　　　
  Takashi FUJISHIRO; Jörg KAHNT; Xiulan XIE; Ulrich ERMLER; Seigo SHIMA
  BMB2015, Dec. 2015, [Domestic conference]
  Japanese, Oral presentation
• Identification of Hcg enzymes in biosynthesis of [Fe]-hydrogenase cofactor　　　　　　　　　　　　　　　
  T. Fujishiro; J. Kahnt; X. Xie; U. Ermler; S. Shima
  ICBIC17, Jul. 2015, [International conference]
  English, Oral presentation
• Structure genomics-based functional identification of Hcg enzymes involved in the biosynthesis of the FeGP cofactor of [Fe]-hydrogenase　　　　　　　　　　　　　　　
  T. Fujishiro; J. Kahnt; X. Xie; U. Ermler; S. Shima
  RIKEN Symposium series: “Metals in Biology, Jun. 2015, [Domestic conference]
  English, Oral presentation
• Functional analysis of HmdII, a [Fe]-hydrogenase paralog, by using its X-ray crystal structure　　　　　　　　　　　　　　　
  T. Fujishiro; K. Ataka; U. Ermler; S. Shima
  The 95th annual meeting of the Chemical Society of Japan, Mar. 2015, [Domestic conference]
  Japanese, Oral presentation
• Structure-guided functional identification of Hcg enzymes involved in the biosynthesis of the [Fe]-hydrogenase bound FeGP cofactor　　　　　　　　　　　　　　　
  T. Fujishiro; J. Kahnt; X. Xie; U. Ermler; S. Shima
  Annual conference 2015 of the association for general and applied microbiology (VAAM-Jahrestagung 2015), Mar. 2015, [International conference]
  English, Poster presentation
• Identification of the HcgB enzyme in biosynthesis of [Fe]-hydrogenase cofactor by a structural genomics-based approach　　　　　　　　　　　　　　　
  T. Fujishiro; H. Tamura; M. Schick; J. Kahnt; X. Xie; K. Ataka; U. Linne; U. Ermler; S. Shima
  EuroBIC12, Aug. 2014, [International conference]
  English, Oral presentation
• Identification of enzymes involved in [Fe]-hydrogenase metal cofactor biosynthesis by a structural genomics-based approach　　　　　　　　　　　　　　　
  T. Fujishiro; H. Tamura; M. Schick; J. Kahnt; X. Xiulan; K. Ataka; U. Ermler; S. Shima
  The 94th annual meeting of the Chemical Society of Japan, Mar. 2014, [Domestic conference]
  Japanese, Oral presentation
• Biochemical and X-ray crystal structure analyses of Hcg proteins involved in biosynthesis of FeGP cofactor of [Fe]-hydrogenase　　　　　　　　　　　　　　　
  T. Fujishiro; H. Tamura; M. Schick; J. Kahnt; X. Xie; U. Ermler; S. Shima
  ICBIC16, Jul. 2013, [International conference]
  English, Poster presentation
• Biochemical analysis of Hcg proteins involved in biosynthesis of the active metallic complex of [Fe]-hydrogenase　　　　　　　　　　　　　　　
  T. Fujishiro; H. Tamura; M. Schick; J. Kahnt; X. Xie; U. Ermler; S. Shima
  The 93rd annual meeting of the Chemical Society of Japan, Mar. 2013, [Domestic conference]
  Japanese, Oral presentation
• Biosynthesis of FeGP cofactor of [Fe]-hydrogenase from methanogenic archaea: functional analyses of an hmd co-occurring gene　　　　　　　　　　　　　　　
  T. Fujishiro; H. Tamura; M. Schick; J. Kahnt; X. Xie; E.Warkentin; U. Ermler; S. Shima
  Gordon research conference of molecular basis of microbial one-carbon metabolism, Aug. 2012, [International conference]
  English, Poster presentation
• Biosynthesis of FeGP cofactor of [Fe]-hydrogenase from methanogenic archaea: functional analyses of an hmd co-occurring gene　　　　　　　　　　　　　　　
  T. Fujishiro; H. Tamura; M. Schick; J. Kahnt; X. Xie; E. Warkentin; U. Ermler; S. Shima
  Gordon research seminar of molecular basis of microbial one-carbon metabolism, Aug. 2012, [International conference]
  English, Poster presentation
• Biochemical analysis of HcgB protein involved in biosynthesis of the active metallic complex of [Fe]-hydrogenase　　　　　　　　　　　　　　　
  T. Fujishiro; H. Tamura; M. Schick; J. Kahnt; X. Xie; U. Ermler; S. Shima
  The 92nd annual meeting of the Chemical Society of Japan, Mar. 2012, [Domestic conference]
  Japanese, Oral presentation
• Structure-based identification of HcgB: implication of guanylyltransfer activity in biosynthesis of the FeGP cofactor of [Fe]-hydrogenase　　　　　　　　　　　　　　　
  T. Fujishiro; H. Tamura; M. Schick; J. Kahnt; X. Xie; E. Warkentin; U. Ermler; S. Shima
  H2-Designcells Meeting 2012, Oct. 2011, [International conference]
  English, Poster presentation
• X-ray crystal structure analysis of hydrogen peroxide-dependent cytochrome P450SPα and the enzymatic oxidation reactions of non-natural substrates　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; T. Watanabe; H. Nakajima; H. Sugimoto; S. Nagano; Y. Shiro; Y. Watanabe
  The 2010 International Chemical Congress of Pacific Basin Societies (Pacifichem2010), Dec. 2010, [International conference]
  English, Poster presentation
• Construction of versatile monooxygenation systems using hydrogen peroxide-dependent cytochrome P450s　　　　　　　　　　　　　　　
  T. Fujishiro
  The 10th Joint Seminar –University of Münster and Nagoya University¬¬–, Nov. 2010, [International conference]
  English, Oral presentation
• Crystal Structure of H2O2-dependent Cytochrome P450SPα from Sphingomonas paucimobilis (CYP152B1)　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; T. Watanabe; H. Sugimoto; S. Nagano; Y. Shiro; Y. Watanabe
  The 5th Asian Biological Inorganic Chemistry Conference (AsBIC V), Nov. 2010, [International conference]
  English, Poster presentation
• X-ray crystal structure analysis of cytochrome P450SPα and enzymatic monooxygenation using hydrogen peroxide　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; T. Watanabe; H. Sugimoto; S. Nagano; Y. Shiro; Y. Watanabe
  The 60th conference of Japan Society of Coordination Chemistry, Sep. 2010, [Domestic conference]
  Japanese, Oral presentation
• X-ray crystal struture analysis of hydrogen peroxide-driven cytochrome P450SPα　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; T. Watanabe; H. Nakajima; H. Sugimoto; S. Nagano; Y. Shiro; Y. Watanabe
  The 10th annual meeting of the Protein Science Society of Japan, Jun. 2010, [Domestic conference]
  Japanese, Poster presentation
• Structural Basis for Understanding Hydrogen Peroxide-dependent Reaction Mechanism of Cytochrome P450SPα　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; T. Watanabe; H. Nakajima; H. Sugimoto; S. Nagano; Y. Shiro; Y. Watanabe
  Nagoya University Grobal COE in Chemistry, Establishment of COE for Elucidation and Design of Materials and Molecular Functions 3rd Annual Symposium, Jun. 2010, [International conference]
  English, Oral presentation
• X-ray crystal structure analysis of cytochrome P450SPα and construction of enzymatic oxidation systems for non-natural substrates　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; T. Watanabe; S. Nagano; Y. Shiro; Y. Watanabe
  The 90th annual meeting of the Chemical Society of Japan, Mar. 2010, [Domestic conference]
  Japanese, Oral presentation
• X-ray Crystal Structure Analysis of Hydrogen Peroxide-dependent Cytochrome P450SPα and Its Application in Oxidation of Non-natural Substrates” The 9th joint seminar University of Münster – Nagoya University　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; T. Watanabe; H. Nakajima; H. Sugimoto; S. Nagano; Y. Shiro; Y. Watanabe
  The 9th joint seminar University of Münster – Nagoya University, Mar. 2010, [International conference]
  English, Oral presentation
• Constructuion of hydrogen peroxide-driven cytochrome P450-based catalytic systems by controlling stereoselectivity with exogenous additives　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; S. Nagano; Y. Shiro; Y. Watanabe
  The 24th symposium on biofunctional chemistry, Sep. 2009, [Domestic conference]
  Japanese, Oral presentation
• Construction of supramolecular enzymatic reaction systems mimicking a substrate-recognition with exogenous additives (decoy molecules) for conversion of stereoselectivity　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; S. Nagano; Y. Shiro; Y. Watanabe
  The summer school of the Japan Society of Coordination Chemistry 2009, Aug. 2009, [Domestic conference]
  Japanese, Poster presentation
• Controlling Enantioselectivity of Enzymatic Sulfoxidation Catalyzed by Cytochrome P450BSβ with Decoy Molecules　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; S. Nagano; Y. Shiro; Y. Watanabe
  ICBIC14, Jul. 2009, [International conference]
  English, Poster presentation
• Regulation mechanism of H2O2 access to the heme cavity in cytochrome P450BSβ　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; S. Tanaka; S. Nagano; Y. Shiro; Y. Watanabe
  16th International Conference on Cytochrome P450, Jun. 2009, [International conference]
  English, Poster presentation
• Controlling the substrate specificity and stereoselectivity of hydrogen peroxide-dependentcytochrome P450 with exogenous additives　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; S. Tanaka; S. Nagano; Y. Shiro; Y. Watanabe
  The 19th symposium on biological reactions involving metals, Jun. 2009, [Domestic conference]
  Japanese, Oral presentation
• Structural changes for substrate- and hydrogen peroxide-binding mechanism of hydrogen peroxide-dependent cytochrome P450　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; S. Tanaka; S. Nagano; Y. Shiro; Y. Watanabe
  The 89th annual meeting of the Chemical Society of Japan, Mar. 2009, [Domestic conference]
  Japanese, Oral presentation
• Structural basis for understanding regulation mechanism of hydrogen peroxide access in the catalytic cycle of cytochrome P450BSβ　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; S. Nagano; Y. Shiro; Y. Watanabe
  Nagoya University Joint Symposia: 5th Yoshimasa Hirata Memorial Lecture and The 1st Global COE International Symposium on Elucidation and Design of Materials and Molecular Functions, Jan. 2009, [International conference]
  English, Poster presentation
• Towards controlling catalytic activity and stereoselectivity of cytochrome P450BSβ with decoy molecules based on its substrate-misrecognition　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; S. Nagano; Y. Shiro; Y. Watanabe
  The 41st symposium on oxidation reactions, Nov. 2008, [Domestic conference]
  Japanese, Oral presentation
• Structural basis for catalytic oxidation of non-natural substrates by cytochrome P450BSβ with decoy molecules　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; H. Nakajima; I. Matsunaga; S. Nagano; Y. Shiro; Y. Watanabe
  The 88th annual meeting of the Chemical Society of Japan, Mar. 2008, [Domestic conference]
  Japanese, Oral presentation
• Construction of a novel biocatalytic system using the substrate misrecognition of cytochrome P450BSβ induced by a decoy molecule　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; H. Nakajima; S. Nagano; Y. Shiro; Y. Watanabe
  Global COE in Chemistry, Nagoya Special Symposium: Featuring the Special Issue of "ACCOUNTS of CHEMICAL RESEARCH": Dioxygen Activation by Metalloenzymes and Models, Mar. 2008, [International conference]
  English, Poster presentation
• Construction of a novel biocatalytic system using the substrate misrecognition of cytochrome P450BSβ induced by a decoy molecule　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; H. Nakajima; I. Matsunaga; S. Nagano; Y. Shiro; Y. Watanabe
  the international conferemce on the research of Organometallic and Cluster Chemistry in Metalloenzymes with Reducing Activities, Mar. 2008, [International conference]
  English, Poster presentation
• Alteration of substrate specificity of heme enzymes using decoy molecules　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; T. Hirose; H. Nakajima; I. Matsunaga; M. Kim; S. Nagano; Y. Shiro; Y. Watanabe
  The 58th conference of Japan Society of Coordination Chemistry, Sep. 2007, [Domestic conference]
  Japanese, Poster presentation
• Construction of a novel biocatalytic system using the substrate misrecognition of heme enzyme　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; T. Hirose; H. Nakajima; I. Matsunaga; M. Kim; S. Nagano; Y. Shiro; Y. Watanabe
  The 2nd International Conference on Joint Project of Chemical Synthesis Core Research Institutions, Aug. 2007, [International conference]
  English, Poster presentation
• Hydrogen peroxide-driven monooxygenation reactions of various non-natural substrates by cytochrome P450BSβ using its substrate misrecognition　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; T. Hirose; H. Nakajima; M. Kim; S. Nagano; Y. Shiro; Y. Watanabe
  The 87th annual meeting of the Chemical Society of Japan, Mar. 2007, [Domestic conference]
  Japanese, Oral presentation
• Addition of different substrate specificities to cytochrome P450BSβ by using substrate analogues　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; H. Nakajima; Y. Shiro; Y. Watanabe
  The 56th conference of Japan Society of Coordination Chemistry, Sep. 2006, [Domestic conference]
  Japanese, Oral presentation
• Formation of active oxidative species used for oxidation of non-natural substrates different from long alkyl-chain-containing fatty acids by hydrogen peroxide-drive cytochrome P450BSβ　　　　　　　　　　　　　　　
  T. Fujishiro; O. Shoji; H. Nakajima; Y. Watanabe
  The 86th annual meeting of the Chemical Society of Japan, Mar. 2006, [Domestic conference]
  Japanese, Oral presentation

• Apr. 2025 - Present
  Basic Biochemistry, Saitama University
• Apr. 2024 - Present
  分子生物学概説
• Sep. 2023 - Present
  Biosynthesis Seminar 2B, Saitama University
• Sep. 2023 - Present
  Biosynthesis Seminar 1B, Saitama University
• Apr. 2023 - Present
  Biosynthesis Seminar 2A, Saitama University
• Apr. 2023 - Present
  Biosynthesis Seminar 1A, Saitama University
• Oct. 2022 - Present
  酵素学
• Apr. 2022 - Present
  基礎生物学実験
• Apr. 2022 - Present
  Metals in Life Science, Saitama University
• Apr. 2021 - Present
  Molecular genetics 7, Saitama University
• Sep. 2020 - Present
  Laboratory work in molecular biological science Ⅱ, Saitama University
• Apr. 2020 - Present
  分子生物科学実験I
• Apr. 2020 - Present
  Seminar in Molecular Cell Biology, Saitama University
• Apr. 2015 - Present
  分子生物学基礎
• Apr. 2015 - Present
  Laboratory Work for Graduation, Saitama University
• Apr. 2022 - Mar. 2023
  生物英語II
• Apr. 2015 - Sep. 2019
  生物英語I

• JAPAN SOCIETY OF COORDINATION CHEMISTRY
• PROTEIN SCIENCE SOCIETY OF JAPAN
• JAPAN SOCIETY FOR BIOSCIENCE, BIOTECHNOLOGY, AND AGROCHEMISTRY
• THE JAPANESE BIOCHEMICAL SOCIETY
• THE CHEMICAL SOCIETY OF JAPAN

• 脱窒とは異なるFe-S型クラスター酵素を中心とした新規な一酸化窒素代謝系の機構解明　　　　　　　　　　　　　　　
  01 Apr. 2025 - 31 Mar. 2028
  Grant amount(Total)：18720000, Direct funding：14400000, Indirect funding：4320000
  Grant number：25K01945
• COを材料にアセチルCoAを合成するニッケル含有酵素複合体の構造と触媒機構解明　　　　　　　　　　　　　　　
  01 Apr. 2025 - 31 Mar. 2027
  Grant amount(Total)：20800000, Direct funding：16000000, Indirect funding：4800000
  Grant number：25H01693
• 三重結合アミノ酸生合成系を改変したクリックケミストリー指向型ペプチド合成系の構築　　　　　　　　　　　　　　　
  01 Apr. 2023 - 31 Mar. 2025
  Grant amount(Total)：10790000, Direct funding：8300000, Indirect funding：2490000
  Grant number：23H04542
  論文ID：49227386, 講演・口頭発表等ID：47281600
• COデヒドロゲナーゼ（CODH）の祖先型酵素HCPがCO代謝活性を得る分子進化の変異の再現　　　　　　　　　　　　　　　
  01 Apr. 2023 - 31 Mar. 2025
  Grant amount(Total)：13000000, Direct funding：10000000, Indirect funding：3000000
  Grant number：23H04648
  論文ID：48150065, 講演・口頭発表等ID：49475338
• Characterization of the unique Fe-S-O metallocluster of HCP enzyme　　　　　　　　　　　　　　　
  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), 01 Apr. 2022 - 31 Mar. 2025
  Saitama University
  Grant amount(Total)：4160000, Direct funding：3200000, Indirect funding：960000
  Grant number：22K05137
  論文ID：43690874, 講演・口頭発表等ID：41945760
• 鉄硫黄クラスター生合成マシナリーの反応過程における過渡的な中間状態の解明　　　　　　　　　　　　　　　
  01 Apr. 2020 - 31 Mar. 2024
  Grant amount(Total)：18070000, Direct funding：13900000, Indirect funding：4170000
  Grant number：20H03204
  講演・口頭発表等ID：41945751
• Unraveling the catalytic mechanisms of Ni-prosthetic-group F430 biosynthetic enzymes　　　　　　　　　　　　　　　
  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area), Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area), 01 Apr. 2019 - 31 Mar. 2021
  Saitama University
  Grant amount(Total)：7540000, Direct funding：5800000, Indirect funding：1740000
  Grant number：19H04639
  論文ID：48322717, 講演・口頭発表等ID：48465474
• Identification of reaction intermediates in iron-sulfur cluster biogenesis for understanding its reaction mechanism　　　　　　　　　　　　　　　
  MEXT, KAKEN(Grant-in-Aid for Young Scientists (B)), Apr. 2017 - Mar. 2020
  Takashi Fujishiro, Principal investigator
  Competitive research funding
  論文ID：48150065, 講演・口頭発表等ID：40155220
• Creation of an artificial hydrogenase enzyme with high activity using in vitro metal cofactor biosynthesis　　　　　　　　　　　　　　　
  MEXT, KAKEN (Grant-in-Aid for Research Activity start-up), Aug. 2015 - Mar. 2017
  Takashi Fujishiro, Principal investigator
  Grant amount(Total)：1430000, Direct funding：1100000, Indirect funding：330000
  Competitive research funding
  書籍等出版物ID：30039801
• フッ素化分子の特異な電子構造を基盤とするヘム酵素の反応空間設計と触媒機能制御　　　　　　　　　　　　　　　
  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for JSPS Fellows, Grant-in-Aid for JSPS Fellows, 2008 - 2010
  Nagoya University
  Grant amount(Total)：1800000, Direct funding：1800000
  Grant number：08J05035

• 20221014 群馬県立前橋女子高等学校（2年生出張講義）　　　　　　　　　　　　　　　
  demonstrator
  14 Oct. 2022 - 14 Oct. 2022