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Biochemistry
電気化学検出の重水素速度論的同位体効果シェワネラ oneidensis氏 1 の細胞外電子輸送
電気化学検出の重水素速度論的同位体効果シェワネラ oneidensis氏 1 の細胞外電子輸送
JoVE Journal
Biochemistry
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JoVE Journal Biochemistry
Electrochemical Detection of Deuterium Kinetic Isotope Effect on Extracellular Electron Transport in Shewanella oneidensis MR-1

電気化学検出の重水素速度論的同位体効果シェワネラ oneidensis氏 1 の細胞外電子輸送

Full Text
10,781 Views
09:00 min
April 16, 2018

DOI: 10.3791/57584-v

Yoshihide Tokunou1, Kazuhito Hashimoto2, Akihiro Okamoto2

1Department of Applied Chemistry,University of Tokyo, 2Global Research Center for Environment and Energy based on Nanomaterials Science,National Institute for Materials Science

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Please note that some of the translations on this page are AI generated. Click here for the English version.

Overview

This protocol outlines whole-cell electrochemical experiments to investigate proton transport's role in extracellular electron transport (EET) in Shewanella oneidensis MR-1. The study employs the deuterium kinetic isotope effect to assess the impact of proton transport on EET kinetics.

Key Study Components

Area of Science

  • Electrochemistry
  • Microbiology
  • Bioenergetics

Background

  • Shewanella oneidensis MR-1 is known for its ability to transfer electrons to electrodes.
  • Proton transport is crucial for the efficiency of extracellular electron transport.
  • The deuterium kinetic isotope effect is a valuable method for studying proton involvement in electron transport.
  • Using living bacteria provides insights into native proton management during EET.

Purpose of Study

  • To observe the contribution of proton transport to the rate of EET.
  • To utilize the deuterium kinetic isotope effect for examining proton transport's role.
  • To improve understanding of the mechanisms underlying bacterial electron transfer.

Methods Used

  • Whole-cell electrochemical experiments.
  • Detection of the deuterium kinetic isotope effect.
  • Use of living Shewanella oneidensis MR-1 cells.
  • Growth of bacterial cultures in LB medium.

Main Results

  • Proton transport significantly affects the kinetics of EET.
  • The deuterium kinetic isotope effect provides clear insights into proton involvement.
  • Noise from other cellular processes was effectively minimized.
  • The method reflects native proton management in EET.

Conclusions

  • Proton transport is a critical factor in the efficiency of EET in bacteria.
  • The protocol offers a reliable approach to study these processes in living cells.
  • Findings contribute to a deeper understanding of microbial electron transfer mechanisms.

Frequently Asked Questions

What is the main focus of this study?
The study focuses on the role of proton transport in extracellular electron transport in Shewanella oneidensis MR-1.
How does the deuterium kinetic isotope effect contribute to the research?
It allows for the examination of proton transport's contribution to electron transport kinetics.
Why use living bacteria instead of purified enzymes?
Using living bacteria reflects the native proton management during electron transport processes.
What challenges were faced during the experiments?
The main challenge was to minimize noise from other cellular processes that could interfere with results.
What are the implications of this research?
The findings enhance the understanding of microbial electron transfer mechanisms and their efficiency.

ここで全細胞外膜シトクロムシェワネラ oneidensis氏-1 複合体を介して細胞外の電子輸送の速度へのプロトン輸送の貢献を研究する電気化学実験のプロトコルを提案する.

このプロトコルの全体的な目標は、重水素速度論的同位体効果の検出により、細菌の細胞外電子輸送速度に対するプロトン輸送の寄与を観察することです。私たちの方法は、プロトン輸送が外膜c型シトクロムを介して細菌から電極への細胞外電子輸送またはEETの速度論に及ぼす影響を明らかにします。私たちのアッセイは、精製された酵素ではなく生きた細菌を使用したため、EETのネイティブプロトン管理を反映しています。

しかし、私たちの課題は、他の細胞プロセスからのノイズを取り除くことでした。KIEと呼ばれる重水素速度論的同位体効果は、関連する電子輸送プロセスに対するプロトン輸送の寄与を調べるための最良の手法の1つです。Shewanella MR-1細胞を増殖させるには、寒天プレート上で増殖させた細胞の1コロニーを15ミリリットルのLB培地に移します。

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