期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 103, 期 19, 页码 7222-7227出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.0511035103
关键词
atomic force microscopy; mechanochemistry
资金
- NHLBI NIH HHS [R01 HL061228, R01 HL066030, HL61228, HL66030] Funding Source: Medline
The mechanism by which mechanical force regulates the kinetics of a chemical reaction is unknown. Here, we use single-molecule force-clamp spectroscopy and protein engineering to study the effect of force on the kinetics of thiol/disulfide exchange. Reduction of disulfide bonds through the thiol/disulfide exchange chemical reaction is crucial in regulating protein function and is known to occur in mechanically stressed proteins. We apply a constant stretching force to single engineered disulfide bonds and measure their rate of reduction by DTT. Although the reduction rate is linearly dependent on the concentration of DTT, it is exponentially dependent on the applied force, increasing 10-fold over a 300-pN range. This result predicts that the disulfide bond lengthens by 0.34 angstrom at the transition state of the thiol/disulfide exchange reaction. Our work at the single bond level directly demonstrates that thiol/disulfide exchange in proteins is a force-dependent chemical reaction. Our findings suggest that mechanical force plays a role in disulfide reduction in vivo, a property that has never been explored by traditional biochemistry. Furthermore, our work also indicates that the kinetics of any chemical reaction that results in bond lengthening will be force-dependent.
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