Conformational changes between the active-site and regulatory light chain of myosin as determined by luminescence resonance energy transfer: The effect of nucleotides and actin

Ming Xiao, Handong Li, Gregory E. Snyder, Roger Cooke, Ralph G. Yount, Paul R. Selvin

Research output: Contribution to journalArticlepeer-review

Abstract

Myosin is thought to generate movement of actin filaments via a conformational change between its light-chain domain and its catalytic domain that is driven by the binding of nucleotides and actin. To monitor this change, we have measured distances between a gizzard regulatory light chain (Cys 108) and the active site (near or at Trp 130) of skeletal myosin subfragment 1 (S1) by using luminescence resonance energy transfer and a photoaffinity ATP-lanthanide analog. The technique allows relatively long distances to be measured, and the label enables site-specific attachment at the active-site with only modest affect on myosin's enzymology. The distance between these sites is 66.8 ± 2.3 Å when the nucleotide is ADP and is unchanged on binding to actin. The distance decreases slightly with ADP- BeF3, (-1.6 ± 0.3 Å) and more significantly with ADP-AIF4 (-4.6 ± 0.2 Å). During steady-state hydrolysis of ATP, the distance is temperature- dependent, becoming shorter as temperature increases and the complex with ADP·P(i) is favored over that with ATP. We conclude that the distance between the active site and the light chain varies as Acto-S1-ADP κ S1-ADP > S1-ADP-BeF3 > S1-ADP-AIF4 ≃ S1-ADP-P(i) and that S1-ATP > S1-ADP-P(i). The changes in distance are consistent with a substantial rotation of the light- chain binding domain of skeletal S1 between the prepowerstroke state, simulated by S1-ADP-AIF4, and the post-powerstroke state, simulated by acto- S1-ADP.

Original languageEnglish (US)
Pages (from-to)15309-15314
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume95
Issue number26
DOIs
StatePublished - Dec 22 1998

ASJC Scopus subject areas

  • General

Fingerprint Dive into the research topics of 'Conformational changes between the active-site and regulatory light chain of myosin as determined by luminescence resonance energy transfer: The effect of nucleotides and actin'. Together they form a unique fingerprint.

Cite this