Catalytic Turnover of [FeFe]-Hydrogenase Based on Single-Molecule Imaging
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- 作者：Christopher Madden ; Michael D. Vaughn ; Ismael D铆ez-P茅rez ; Katherine A. Brown ; Paul W. King ; Devens Gust ; Ana L. Moore ; Thomas A. Moore
- 刊名：The Journal of the American Chemical Society
- 出版年：2012
- 出版时间：January 25, 2012
- 年：2012
- 卷：134
- 期：3
- 页码：1577-1582
- 全文大小：872K
- 年卷期：v.134,no.3(January 25, 2012)
- ISSN：1520-5126
文摘
Hydrogenases catalyze the interconversion of protons and hydrogen according to the reversible reaction: 2H+ + 2e鈥?/sup> H2 while using only the earth-abundant metals nickel and/or iron for catalysis. Due to their high activity for proton reduction and the technological significance of the H+/H2 half reaction, it is important to characterize the catalytic activity of [FeFe]-hydrogenases using both biochemical and electrochemical techniques. Following a detailed electrochemical and photoelectrochemical study of an [FeFe]-hydrogenase from Clostridium acetobutylicum (CaHydA), we now report electrochemical and single-molecule imaging studies carried out on a catalytically active hydrogenase preparation. The enzyme CaHydA, a homologue (70% identity) of the [FeFe]-hydrogenase from Clostridium pasteurianum, CpI, was adsorbed to a negatively charged, self-assembled monolayer (SAM) for investigation by electrochemical scanning tunneling microscopy (EC-STM) techniques and macroscopic electrochemical measurements. The EC-STM imaging revealed uniform surface coverage with sufficient stability to undergo repeated scanning with a STM tip as well as other electrochemical investigations. Cyclic voltammetry yielded a characteristic cathodic hydrogen production signal when the potential was scanned sufficiently negative. The direct observation of the single enzyme distribution on the Au-SAM surface coupled with macroscopic electrochemical measurements obtained from the same electrode allowed the evaluation of a turnover frequency (TOF) as a function of potential for single [FeFe]-hydrogenase molecules.