YUZAWA Satoshi

写真a

Affiliation

Graduate School of Media and Governance Institute for Advanced Biosciences ( Shonan Fujisawa )

Position

Project Senior Assistant Professor (Non-tenured)/Project Assistant Professor (Non-tenured)/Project Lecturer (Non-tenured)

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External Links
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Profile Summary 【 Display / hide 】

  • 20世紀の化学合成は肥料や医薬品を生み出し社会の繁栄を支えましたが、現代は気候変動や地政学リスクに伴うサプライチェーンの脆弱性など新たな課題に直面し、従来の化石資源依存のモノづくりは限界を迎えています。

    湯澤ラボは、化学合成の制約を超える「合成生物学」でこれらの課題に挑みます。特許出願済みの長鎖DNA合成・編集技術で巨大酵素と微生物を高度に制御し、化学的手法では製造困難な多種多様な高付加価値分子（分子量100-1000）を、持続可能かつ自立したプロセスで生産する技術を確立します。

    高度な分子製造技術を日本の強みとし、複雑な機能性分子を自在に創出することで、経済安全保障を高め、不確実な時代でも揺るがない強靭で豊かな社会の実現に貢献します。

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Books 【 Display / hide 】

  • バイオエネルギー再燃

    植田, 充美, シーエムシー出版, 2021.11,  Page： vi, 289p

  • Comprehensive Natural Products III

    Hung-Wen (Ben) Liu and Tadhg P. Begley, Elsevier, 2020

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Papers 【 Display / hide 】

  • Extensive Alanine Scanning of Loop Regions in Ketosynthase Domains Identifies Non-Active Site Mutations with Drastic Effects on Polyketide Biosynthesis.

    Hiromitsu Yamamoto, Hiroko Ueda, Misaki Aso, Minjae Lee, Satoshi Yuzawa

    ACS chemical biology 21 ( 2 ) 362 - 370 2026.02

    ISSN  15548929

    　View Summary

    Modular polyketide synthases (PKSs) produce diverse natural products with significant pharmaceutical value, but their protein engineering for drug discovery is hampered by the unpredictable substrate specificity of their ketosynthase (KS) domains. While previous studies have focused on the KS active sites, we conducted extensive alanine scanning of loop regions of a KS domain, most of which are distant from the catalytic center. In vitro screening of 46 point mutants revealed that ∼70% of the mutants retained their activity, whereas ∼25% of the mutants displayed severely reduced activity, and two mutants unexpectedly showed enhanced activity. Interestingly, most mutations with significant impact were located more than 20 Å away from the catalytic center. These findings provide the first clear evidence that KS residues beyond the canonical active site play crucial roles in polyketide biosynthesis. Our results show essential foundational data to understand KS functions, which could be used for developing more effective KS engineering strategies beyond traditional active site modifications.

  • Direct Pathway Synthesis and Editing (DiPaSE): A One-Pot DNA Assembly Method for Accurate and Efficient Refactoring of High-GC Biosynthetic Gene Clusters.

    Tomoki Takeda, Misaki Aso, Hiroko Ueda, Satoshi Yuzawa

    ACS synthetic biology 15 ( 3 ) 1221 - 1230 2026.02

    　View Summary

    Natural products (NPs) produced by actinobacteria, particularly Streptomyces species, represent a rich source of bioactive compounds and have yielded many clinically important compounds. Actinobacterial genomes are characterized by high GC content and typically harbor 20-40 biosynthetic gene clusters (BGCs) per genome, which encode diverse NPs such as polyketides, peptides, and glycosides. CRISPR/Cas-based genome editing has emerged as a promising tool to activate silent BGCs and engineer NP biosynthesis. However, the efficiency of multiplex editing drastically decreases as the number of targeted sites increases. Here, we report a novel one-pot DNA assembly method, termed direct pathway synthesis and editing (DiPaSE), for the efficient synthesis and multiplex editing of long, high-GC BGCs. DiPaSE accurately assembles multiple high-GC DNA fragments up to 60 kb and enables simultaneous deletions and insertions within a target BGC without compromising the assembly efficiency. Using this approach, we identified functions of previously uncharacterized genes in the aureothin BGC and significantly enhanced the titer of the corresponding NP. The workflow employs conventional polymerase chain reaction, type IIP restriction enzymes, commercially available DNA assembly reagents, and Escherichia coli, providing a simple, cost-effective, and broadly applicable platform for genome mining, BGC refactoring, and rational design of artificial biosynthetic pathways.

  • Engineering Pseudomonas putida for production of 3-hydroxyacids using hybrid type I polyketide synthases.

    Matthias Schmidt, Aaron A Vilchez, Namil Lee, Leah S Keiser, Allison N Pearson, Mitchell G Thompson, Yolanda Zhu, Robert W Haushalter, Adam M Deutschbauer, Satoshi Yuzawa, Lars M Blank, Jay D Keasling

    Metabolic engineering communications 20   e00261 2025.06

    　View Summary

    Engineered type I polyketide synthases (T1PKSs) are a potentially transformative platform for the biosynthesis of small molecules. Due to their modular nature, T1PKSs can be rationally designed to produce a wide range of bulk or specialty chemicals. While heterologous PKS expression is best studied in microbes of the genus Streptomyces, recent studies have focused on the exploration of non-native PKS hosts. The biotechnological production of chemicals in fast growing and industrial relevant hosts has numerous economic and logistic advantages. With its native ability to utilize alternative feedstocks, Pseudomonas putida has emerged as a promising workhorse for the sustainable production of small molecules. Here, we outline the assessment of P. putida as a host for the expression of engineered T1PKSs and production of 3-hydroxyacids. After establishing the functional expression of an engineered T1PKS, we successfully expanded and increased the pool of available acyl-CoAs needed for the synthesis of polyketides using transposon sequencing and protein degradation tagging. This work demonstrates the potential of T1PKSs in P. putida as a production platform for the sustainable biosynthesis of unnatural polyketides.

  • Identification of Key Amino Acids in the A Domains of Polymyxin Synthetase Responsible for 2,4-Diaminobutyric Acid Adenylation in Paenibacillus polymyxa NBRC3020 Strain

    Mai Nemoto, Wataru Ando, Taichi Mano, Minjae Lee, Satoshi Yuzawa, Toshihisa Mizuno

    ACS Chemical Biology 20 ( 2 ) 321 - 331 2025.01

    Accepted,  ISSN  15548929

    　View Summary

    Developing novel nonribosomal peptides (NRPs) requires a comprehensive understanding of the enzymes involved in their biosynthesis, particularly the substrate amino acid recognition mechanisms in the adenylation (A) domain. This study focused on the A domain responsible for adenylating l-2,4-diaminobutyric acid (l-Dab) within the synthetase of polymyxin, an NRP produced by Paenibacillus polymyxa NBRC3020. To date, investigations into recombinant proteins that selectively adenylate l-Dab─exploring substrate specificity and enzymatic activity parameters─have been limited to reports on A domains found in enzymes synthesizing l-Dab homopolymers (pldA from S. celluloflavus USE31 and pddA from S. hindustanus NBRC15115), which remain exceedingly rare. The polymyxin synthetase in NBRC3020 contains five A domains specific to l-Dab, distributed across five distinct modules (modules 1, 3, 4, 5, 8, and 9). In this study, we successfully obtained soluble A domain proteins from modules 1, 5, 8, and 9 by preparing module-specific recombinant proteins. These proteins were expressed in E. coli BAP-1, purified via Ni-affinity chromatography, and demonstrated high specificity for l-Dab. Through sequence homology analysis, three-dimensional structural modeling, docking simulations to estimate substrate-binding sites, and functional validation using alanine mutants, we identified Glu281 and Asp344 as critical residues for recognizing the side chain amino group of l-Dab, and Asp238 as essential for recognizing its main chain amino group in the A domain. Notably, these key residues were conserved not only across the A domains in modules 1, 5, 8, and 9 of P. polymyxa NBRC3020 but also in those of the P. polymyxa PKB1 strain, as confirmed by sequence homology analysis. Interestingly, in pldA and pddA, the key residues involved in recognizing the side-chain amino group of l-Dab, which are conserved among polymyxin synthetases of NBRC3020 and PKB1 strain, were not observed. This suggests a potentially different mechanism for l-Dab recognition.

  • Genome sequences of three Streptomyces isolated from the soil of soybean field in Tsuruoka, Japan.

    Ryuki Sato, Rena Saito, Tomoki Takeda, Nobuhiro Sasaki, Satoshi Yuzawa, Natsumi Saito, Kazuharu Arakawa

    Microbiology resource announcements 14 ( 1 ) e0105924 2025.01

    　View Summary

    Streptomyces are Gram-positive bacteria typically found in the soil, with very large genomes and high GC content, and are known to produce a wide range of secondary metabolites. We isolated and sequenced the genomes of three Streptomyces bacteria from the soil of soybean field in Tsuruoka, Japan.

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Papers, etc., Registered in KOARA 【 Display / hide 】

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Reviews, Commentaries, etc. 【 Display / hide 】

  • ガソリンの微生物生産に向けた合成生物学的アプローチ

    湯澤 賢

    バイオサイエンスとインダストリー (バイオインダストリー協会)  77 ( 6 ) 440 - 443 2019

    ISSN  0914-8981

  • Interview (2)(就職支援OG・OBインタビュー編,バイオ系のキャリアデザイン)

    湯澤 賢

    生物工学会誌 : seibutsu-kogaku kaishi (日本生物工学会)  93 ( 12 ) 767 - 768 2015

    ISSN  0919-3758

  • 研究コミュニティーとつながる連載 研コミュ白書(第5回)スタンフォードLSJ(Life Science in Japanese) : アメリカ西海岸の日本人研究者コミュニティー

    藤島 皓介, 湯澤 賢

    細胞工学 (学研メディカル秀潤社 ; 1982-)  32 ( 3 ) 344 - 349 2013

    ISSN  0287-3796

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Research Projects of Competitive Funds, etc. 【 Display / hide 】

  • 人工PKSを用いた次世代バイオ燃料の生産

    2026.04
    -
    2028.03

    科学技術振興機構, 革新的GX技術創出事業, No Setting

  • 合成生物学的手法による液体燃料の自在合成基盤の確立

    2022.04
    -
    2024.03

    公益財団法人・発酵研究所, 一般研究助成, Research grant, Principal investigator

  • 非大腸菌タンパク質生産系を併用したモジュラーポリケチド合成酵素の試験管内再構成

    2021.04
    -
    2024.03

    MEXT,JSPS, Grant-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), Principal investigator

    　View Summary

    モジュラーポリケチド合成酵素（モジュラーPKS）は多様な薬剤の基本骨格の生合成を担う巨大なマルチドメイン酵素であり、その機能や構造の解析が盛んに試みられている。しかしタンパク質の分割など大規模な改変を加えていない野生型酵素による試験管内再構成を達成した例は世界でまだ1例しかなく、至適温度、至適pH、至適イオン強度、kcat、KM等の基本的な情報が一般にどんな分布を示すかは明らかになっていない。本研究では、これまで利用されていない非大腸菌タンパク質生産系も併用し、新たに数種類の野生型モジュラーPKSの試験管内再構成およびそれらの生化学的解析を実施する。当該酵素の理解がより一層進めば、モジュラーPKSの機能改変による医薬品アナログ等の開発も飛躍的に進展すると考えられる。第一のモジュラーPKSに関しては、非大腸菌タンパク質生産系を併用することで試験管内再構成にすでに成功している（通常の大腸菌を用いたタンパク質生産系では酵素の活性体を一部取得できなかった）。また、至適温度、至適pH、至適イオン強度、kcat、KM等の解析を現在進めているところである。本成果は未発表であるが、世界で2例目、国内では初の成果となり得る。第二のモジュラーPKSに関しては、プラスミド構築やタンパク質精製など試験管内再構成に向けた準備を行っている段階である。また、本研究計画提出時には記載していなかったが、現在第三の系の再構成の準備も進めている。この第三の系は、モジュラーPKSではなく、モジュラーPKSと非リボソーム型ペプチド合成酵素とのハイブリッドの系である。予備実験の結果、大腸菌を用いた通常のタンパク質生産系では一部酵素の活性体の取得が困難であることが明らかとなった。今後は、我々が保有する非大腸菌タンパク質生産系を活用して、試験管内再構成および生化学的解析を実施する予定である。

  • 合成生物学的手法による抗生物質の自在合成基盤の確立

    2021.04
    -
    2024.03

    文部科学省・科学技術振興機構, 創発的研究支援事業（フェーズ１）, No Setting, Principal investigator

  • Structural analysis of a modular polyketide synthase module

    2019.08
    -
    2021.03

    MEXT,JSPS, Grant-in-Aid for Scientific Research, YUZAWA Satoshi, Grant-in-Aid for Research Activity Start-up , Principal investigator

    　View Summary

    Here we proposed to characterize a protein shape(s) of modular polyketide synthases using small-angle X-ray scattering (SAXS), cryo-electron microscopy, and a biochemical assay that has not been applied to modular PKSs. Although biochemical characterization of the structure is still ongoing, our SAXS data indicates that each module of modular PKSs forms a X-shape structure, which challenges the current arched shape model reported in Nature. We will continue to analyze a PKS structure to provide an accurate structural model to our research community.

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Courses Taught 【 Display / hide 】

  • GENETIC ANALYSIS LABORATORY

    2026

  • GENETIC ANALYSIS LABORATORY

    2025

  • GENETIC ANALYSIS LABORATORY

    2024

  • GENETIC ANALYSIS LABORATORY

    2023

  • GENETIC ANALYSIS LABORATORY

    2022

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