Oct 31, 2018

High-resolution models of actin-bound myosin from EPR of a bifunctional spin label

BioRxiv : the Preprint Server for Biology
Benjamin P. BinderDavid D. Thomas

Abstract

We have employed two complementary high-resolution electron paramagnetic resonance (EPR) techniques with a bifunctional spin label (BSL) to test and refine protein structural models based on crystal structures and cryo-EM. We demonstrate this approach by investigating the effects of nucleotide binding on the structure of myosin's catalytic domain (CD), while myosin is in complex with actin. Unlike conventional spin labels attached to single Cys, BSL reacts with a pair of Cys; in this study, we thoroughly characterize BSL's rigid, highly stereoselective attachment to protein α-helices, which permits accurate measurements of orientation and distance. Distance constraints were obtained from double electron-electron resonance (DEER) on myosin constructs labeled with BSL specifically at two sites. Constraints for orientation of individual helices were obtained previously from continuous-wave EPR (CW-EPR) of myosin labeled at specific sites with BSL in oriented muscle fibers. We have shown previously that CW-EPR of BSL quantifies helix orientation within actin-bound myosin; here we show that the addition of high-resolution distance constraints by DEER alleviates remaining spatial ambiguity, allowing for direct testing and refinement ...Continue Reading

  • References
  • Citations

References

  • We're still populating references for this paper, please check back later.
  • References
  • Citations

Citations

  • This paper may not have been cited yet.

Mentioned in this Paper

Study
Muscle Rigidity
Tissue Membrane
Molecular Helix
Membrane
Actins
Nucleotide Binding
Spatial Distribution
Filamentous fungus
Helix (Snails)

About this Paper

Related Feeds

BioRxiv & MedRxiv Preprints

BioRxiv and MedRxiv are the preprint servers for biology and health sciences respectively, operated by Cold Spring Harbor Laboratory. Here are the latest preprint articles (which are not peer-reviewed) from BioRxiv and MedRxiv.