Atomistic Models from Orientation and Distance Constraints Using EPR of a Bifunctional Spin Label

Benjamin P. Binder, Andrew R Thompson, David D Thomas

Research output: Contribution to journalArticle

Abstract

We have used high-resolution orientation and distance measurements derived from electron paramagnetic resonance of a bifunctional spin label (BSL) to build and refine atomistic models of protein structure. We demonstrate this approach by investigating the effects of nucleotide binding on the structure of myosin's catalytic domain while myosin is in complex with actin. Constraints for orientation of individual helices were obtained in a previous study from continuous-wave electron paramagnetic resonance of myosin labeled at specific sites with BSLs in oriented muscle fibers. In this study, new distance constraints were derived from double electron-electron resonance on myosin constructs labeled with a BSL specifically at two sites. Using these complementary constraints together, we thoroughly characterize the BSL's rigid, highly stereoselective attachment to protein α-helices, which permits accurate measurements of orientation and distance. We also leverage these measurements to derive a novel, to our knowledge, structural model for myosin-II in complex with actin and MgADP and compare our model to other recent actomyosin structures. The described approach is applicable to any orientable complex (e.g., membranes or filaments) in which site-specific di-Cys mutation is feasible.

Original languageEnglish (US)
Pages (from-to)319-330
Number of pages12
JournalBiophysical journal
Volume117
Issue number2
DOIs
StatePublished - Jul 23 2019

Fingerprint

Spin Labels
Myosins
Electron Spin Resonance Spectroscopy
Actins
Electrons
Myosin Type II
Actomyosin
Structural Models
Adenosine Diphosphate
Catalytic Domain
Proteins
Nucleotides
Muscles
Mutation
Membranes

PubMed: MeSH publication types

  • Journal Article

Cite this

Atomistic Models from Orientation and Distance Constraints Using EPR of a Bifunctional Spin Label. / Binder, Benjamin P.; Thompson, Andrew R; Thomas, David D.

In: Biophysical journal, Vol. 117, No. 2, 23.07.2019, p. 319-330.

Research output: Contribution to journalArticle

@article{0f4f594b0fc74cbc9a34bbd1909dabb7,
title = "Atomistic Models from Orientation and Distance Constraints Using EPR of a Bifunctional Spin Label",
abstract = "We have used high-resolution orientation and distance measurements derived from electron paramagnetic resonance of a bifunctional spin label (BSL) to build and refine atomistic models of protein structure. We demonstrate this approach by investigating the effects of nucleotide binding on the structure of myosin's catalytic domain while myosin is in complex with actin. Constraints for orientation of individual helices were obtained in a previous study from continuous-wave electron paramagnetic resonance of myosin labeled at specific sites with BSLs in oriented muscle fibers. In this study, new distance constraints were derived from double electron-electron resonance on myosin constructs labeled with a BSL specifically at two sites. Using these complementary constraints together, we thoroughly characterize the BSL's rigid, highly stereoselective attachment to protein α-helices, which permits accurate measurements of orientation and distance. We also leverage these measurements to derive a novel, to our knowledge, structural model for myosin-II in complex with actin and MgADP and compare our model to other recent actomyosin structures. The described approach is applicable to any orientable complex (e.g., membranes or filaments) in which site-specific di-Cys mutation is feasible.",
author = "Binder, {Benjamin P.} and Thompson, {Andrew R} and Thomas, {David D}",
year = "2019",
month = "7",
day = "23",
doi = "10.1016/j.bpj.2019.04.042",
language = "English (US)",
volume = "117",
pages = "319--330",
journal = "Biophysical Journal",
issn = "0006-3495",
publisher = "Biophysical Society",
number = "2",

}

TY - JOUR

T1 - Atomistic Models from Orientation and Distance Constraints Using EPR of a Bifunctional Spin Label

AU - Binder, Benjamin P.

AU - Thompson, Andrew R

AU - Thomas, David D

PY - 2019/7/23

Y1 - 2019/7/23

N2 - We have used high-resolution orientation and distance measurements derived from electron paramagnetic resonance of a bifunctional spin label (BSL) to build and refine atomistic models of protein structure. We demonstrate this approach by investigating the effects of nucleotide binding on the structure of myosin's catalytic domain while myosin is in complex with actin. Constraints for orientation of individual helices were obtained in a previous study from continuous-wave electron paramagnetic resonance of myosin labeled at specific sites with BSLs in oriented muscle fibers. In this study, new distance constraints were derived from double electron-electron resonance on myosin constructs labeled with a BSL specifically at two sites. Using these complementary constraints together, we thoroughly characterize the BSL's rigid, highly stereoselective attachment to protein α-helices, which permits accurate measurements of orientation and distance. We also leverage these measurements to derive a novel, to our knowledge, structural model for myosin-II in complex with actin and MgADP and compare our model to other recent actomyosin structures. The described approach is applicable to any orientable complex (e.g., membranes or filaments) in which site-specific di-Cys mutation is feasible.

AB - We have used high-resolution orientation and distance measurements derived from electron paramagnetic resonance of a bifunctional spin label (BSL) to build and refine atomistic models of protein structure. We demonstrate this approach by investigating the effects of nucleotide binding on the structure of myosin's catalytic domain while myosin is in complex with actin. Constraints for orientation of individual helices were obtained in a previous study from continuous-wave electron paramagnetic resonance of myosin labeled at specific sites with BSLs in oriented muscle fibers. In this study, new distance constraints were derived from double electron-electron resonance on myosin constructs labeled with a BSL specifically at two sites. Using these complementary constraints together, we thoroughly characterize the BSL's rigid, highly stereoselective attachment to protein α-helices, which permits accurate measurements of orientation and distance. We also leverage these measurements to derive a novel, to our knowledge, structural model for myosin-II in complex with actin and MgADP and compare our model to other recent actomyosin structures. The described approach is applicable to any orientable complex (e.g., membranes or filaments) in which site-specific di-Cys mutation is feasible.

UR - http://www.scopus.com/inward/record.url?scp=85068527386&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85068527386&partnerID=8YFLogxK

U2 - 10.1016/j.bpj.2019.04.042

DO - 10.1016/j.bpj.2019.04.042

M3 - Article

C2 - 31301803

AN - SCOPUS:85068527386

VL - 117

SP - 319

EP - 330

JO - Biophysical Journal

JF - Biophysical Journal

SN - 0006-3495

IS - 2

ER -