One at a Time: Intramolecular Electron-Transfer Kinetics in Small Laccase Observed during Turnover
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- 作者：Ankur Gupta ; Thijs J. Aartsma ; Gerard W. Canters
- 刊名：Journal of the American Chemical Society
- 出版年：2014
- 出版时间：February 19, 2014
- 年：2014
- 卷：136
- 期：7
- 页码：2707-2710
- 全文大小：258K
- ISSN：1520-5126
文摘
Single-molecule enzymology provides an unprecedented level of detail about aspects of enzyme mechanisms which have been very difficult to probe in bulk. One such aspect is intramolecular electron transfer (ET), which is a recurring theme in the research on oxidoreductases containing multiple redox-active sites. We measure the intramolecular ET rates between the copper centers of the small laccase from Streptomyces coelicolor at room temperature and pH 7.4, one molecule at a time, during turnover. The forward and backward rates across many molecules follow a log-normal distribution with means of 460 and 85 s鈥?, respectively, corresponding to activation energies of 347 and 390 meV for the forward and backward rates. The driving force and the reorganization energy amount to 0.043 and 1.5 eV, respectively. The spread in rates corresponds to a spread of 30 meV in the activation energy. The second-order rate constant for reduction of the T1 site amounts to 2.9 脳 104 M鈥? s鈥?. The mean of the distribution of forward ET rates is higher than the turnover rate from ensemble steady-state measurements and, thus, is not rate limiting.