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Nanoscale probing and visualization of cyclic nucleotide-gated ion channels structural dynamics and function

Research Project

Project/Area Number 20K22626
Research Category

Grant-in-Aid for Research Activity Start-up

Allocation TypeMulti-year Fund
Review Section 0701:Biology at molecular to cellular levels, and related fields
Research InstitutionKanazawa University

Principal Investigator

Marchesi Arin  金沢大学, ナノ生命科学研究所, 特任助教 (80882240)

Project Period (FY) 2020-09-11 – 2022-03-31
Project Status Discontinued (Fiscal Year 2021)
Budget Amount *help
¥2,860,000 (Direct Cost: ¥2,200,000、Indirect Cost: ¥660,000)
Fiscal Year 2021: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2020: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Keywordsion channels / gating / atomic force microscopy / membrane proteins / CNG channels / Atomic Force Microscopy / Ion channels / Purification / Gating
Outline of Research at the Start

The proposal will advance our understanding of ion channels function by exploiting a recently developed mechanical microscope in combination to advanced optics. The combination of these techniques will allow us to study ion channels structural dynamcs and link it to their physiological function.

Outline of Annual Research Achievements

The SthK channel gene (UniProtKB G0GA88)was subcloned in the appropriate vector (pCGFP-BC) for overexpression in bacteria (BL21-DE3). Channel expression, purification and reconstitution in artificial bilayer was implemented successfully. The sample was characterized by means of force-spectroscopy (i.e. adsorbed to the AFM probe and unfolded from the membrane bilayer) and high-speed AFM imaging both in the presence of the endogenous ligand (cAMP) and its absence. This allowed us to define key interactions among the different protein domains and the overall mechanisms of gating (i.e. the opening of the permeation pathway). One paper, in collaboration with the ISAS-Italy (International School for Advanced Studies) is currently in revision and another is in preparation.

Report

(2 results)
  • 2021 Annual Research Report
  • 2020 Research-status Report
  • Research Products

    (4 results)

All 2021 Other

All Int'l Joint Research (2 results) Journal Article (2 results) (of which Int'l Joint Research: 2 results,  Peer Reviewed: 2 results,  Open Access: 1 results)

  • [Int'l Joint Research] SISSA/ISAS(イタリア)

    • Related Report
      2021 Annual Research Report
  • [Int'l Joint Research] SISSA/ISAS(イタリア)

    • Related Report
      2020 Research-status Report
  • [Journal Article] CNG channel structure, function, and gating: a tale of conformational flexibility2021

    • Author(s)
      Napolitano Luisa Maria Rosaria、Torre Vincent、Marchesi Arin
    • Journal Title

      Pfl?gers Archiv - European Journal of Physiology

      Volume: 473 Issue: 9 Pages: 1423-1435

    • DOI

      10.1007/s00424-021-02610-6

    • Related Report
      2021 Annual Research Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] An ultra-wide scanner for large-area high-speed atomic force microscopy with megapixel resolution2021

    • Author(s)
      Marchesi Arin、Umeda Kenichi、Komekawa Takumi、Matsubara Takeru、Flechsig Holger、Ando Toshio、Watanabe Shinji、Kodera Noriyuki、Franz Clemens M.
    • Journal Title

      Scientific Reports

      Volume: 11 Issue: 1 Pages: 13003-13003

    • DOI

      10.1038/s41598-021-92365-y

    • Related Report
      2021 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research

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Published: 2020-09-29   Modified: 2022-12-28  

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