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非天然型アミノ酸生産のためのPLP依存性アミノ基転移酵素の改変

Research Project

Project/Area Number 19F19764
Research Category

Grant-in-Aid for JSPS Fellows

Allocation TypeSingle-year Grants
Section外国
Review Section Basic Section 43020:Structural biochemistry-related
Research InstitutionOkinawa Institute of Science and Technology Graduate University

Principal Investigator

LAURINO Paola  沖縄科学技術大学院大学, タンパク質工学・進化ユニット, 准教授 (90812256)

Co-Investigator(Kenkyū-buntansha) DINDO MIRCO  沖縄科学技術大学院大学, タンパク質工学・進化ユニット, 外国人特別研究員
Project Period (FY) 2019-11-08 – 2022-03-31
Project Status Completed (Fiscal Year 2021)
Budget Amount *help
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 2021: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2020: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 2019: ¥100,000 (Direct Cost: ¥100,000)
KeywordsPhase separation / enzymes / enzyme kinetic / protein engineering / protein plasticity / Disordered regions / AGT / protocells / Liquid phase separation / Biochemistry / enzyme / library / screening
Outline of Research at the Start

The DPAT enzyme will be fully characterised from structural and functional perspectives, in order to understand its unusual activity in detail. Laurino group already has an established collaboration with the Jackson Lab at ANU, Australia and they will help to solve the structure of this enzyme. Then, BLAST searches will help us to find proteins structurally related to DPAT, in order to identify possible homologs with the same or similar functions and to expand our knowledge of this interesting class of enzymes. Finally the recombinant DPAT will be engineered by rounds of directed evolution.

Outline of Annual Research Achievements

・The production/generation of liquid-liquid phase separated membrane-less droplets with novel features as the ability to compartmentalize enzymes efficiently, high protein crowding (mM protein content) to study in detail the effects of crowding on proteins and especially on the enzymatic activity (using length scale of living systems micrometer scale as cells).
・We also demonstrated how the enzymatic activity in presence of crowding generated flow similar to the cytoplasmatic streaming of the cell. This work has been published by Nature Communications it is amongst the 2021 top 25 most downloaded Chemistry and Material Science Articles in 2021 published in Nature communication.
・In addition, we have discovered how protein plasticity shape the function and fitness. The work has been published in Protein Science.

Research Progress Status

令和3年度が最終年度であるため、記入しない。

Strategy for Future Research Activity

令和3年度が最終年度であるため、記入しない。

Report

(3 results)
  • 2021 Annual Research Report
  • 2020 Annual Research Report
  • 2019 Annual Research Report
  • Research Products

    (4 results)

All 2022 2021 2020 2019

All Journal Article (4 results) (of which Int'l Joint Research: 4 results,  Peer Reviewed: 4 results,  Open Access: 4 results)

  • [Journal Article] Structural dynamics shape the fitness window of alanine:glyoxylate aminotransferase2022

    • Author(s)
      Dindo Mirco、Pascarelli Stefano、Chiasserini Davide、Grottelli Silvia、Costantini Claudio、Uechi Gen‐Ichiro、Giardina Giorgio、Laurino Paola、Cellini Barbara
    • Journal Title

      Protein Science

      Volume: 31 Issue: 5 Pages: 1-10

    • DOI

      10.1002/pro.4303

    • Related Report
      2021 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Sustained enzymatic activity and flow in crowded protein droplets2021

    • Author(s)
      Testa Andrea、Dindo Mirco、Rebane Aleksander A.、Nasouri Babak、Style Robert W.、Golestanian Ramin、Dufresne Eric R.、Laurino Paola
    • Journal Title

      Nature Communications

      Volume: 12 Issue: 1 Pages: 1-10

    • DOI

      10.1038/s41467-021-26532-0

    • Related Report
      2021 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Non-enzymatic and highly sensitive lactose detection utilizing graphene field-effect transistors2020

    • Author(s)
      Danielson Eric、Dindo Mirco、Porkovich Alexander J.、Kumar Pawan、Wang Zhenwei、Jain Prashant、Mete Trimbak、Ziadi Zakaria、Kikkeri Raghavendra、Laurino Paola、Sowwan Mukhles
    • Journal Title

      Biosensors and Bioelectronics

      Volume: 165 Pages: 112419-112419

    • DOI

      10.1016/j.bios.2020.112419

    • Related Report
      2020 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Cycloserine enantiomers are reversible inhibitors of human alanine:glyoxylate aminotransferase: implications for Primary Hyperoxaluria type 12019

    • Author(s)
      Mirco Dindo; Silvia Grottelli; Giannamaria Annunziato; Giorgio Giardina; Marco Pieroni; Gioena Pampalone; Andrea Faccini; Francesca Cutruzzola; Paola Laurino; Gabriele Costantino; Barbara Cellini
    • Journal Title

      Biochem J

      Volume: 476 Issue: 24 Pages: 3751-3768

    • DOI

      10.1042/bcj20190507

    • Related Report
      2019 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research

URL: 

Published: 2019-11-29   Modified: 2024-03-26  

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