Format

Send to

Choose Destination
Nat Methods. 2012 Oct;9(10):1013-9. doi: 10.1038/nmeth.2152. Epub 2012 Sep 2.

Ultrafast force-clamp spectroscopy of single molecules reveals load dependence of myosin working stroke.

Author information

1
European Laboratory for Non-linear Spectroscopy, University of Florence, Sesto Fiorentino, Italy. capitan@lens.unifi.it

Abstract

We describe a dual-trap force-clamp configuration that applies constant loads between a binding protein and an intermittently interacting biological polymer. The method has a measurement delay of only ∼10 μs, allows detection of interactions as brief as ∼100 μs and probes sub-nanometer conformational changes with a time resolution of tens of microseconds. We tested our method on molecular motors and DNA-binding proteins. We could apply constant loads to a single motor domain of myosin before its working stroke was initiated (0.2-1 ms), thus directly measuring its load dependence. We found that, depending on the applied load, myosin weakly interacted (<1 ms) with actin without production of movement, fully developed its working stroke or prematurely detached (<5 ms), thus reducing the working stroke size with load. Our technique extends single-molecule force-clamp spectroscopy and opens new avenues for investigating the effects of forces on biological processes.

PMID:
22941363
DOI:
10.1038/nmeth.2152
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Nature Publishing Group
Loading ...
Support Center