Visualization of Domains in Native and Nucleotide-trapped Myosin Heads by Negative Staining

Overview

Journal J Muscle Res Cell Motil

Specialties Cell Biology
Physiology

Date 1988 Aug 1

PMID 2464615

Citations 13

Authors

M Walker

J Trinick

Affiliations

Soon will be listed here.

Abstract

Electron microscopy of negatively stained vertebrate skeletal muscle myosin molecules has revealed substructure suggestive of globular domains in the head portions of the molecule. This head substructure has been examined after both low and high electron doe. The results suggest it is probably not an artefact of radiation damage. The most common appearance is of one or two stain-filled clefts which run roughly perpendicular to the long axis of the head, giving rise to the appearance of two or three domains in a line. A large domain is located at the end of the head, while two smaller domains are arranged between this and the head-tail junction. The size of the large distal domain (about 10 nm long and about 7 nm wide at its widest point) is similar in heads showing either two or three domains. Stable analogues of M.ATP and M.ADP.Pi, the predominant complexes present during hydrolysis of ATP by myosin, were prepared by crosslinking the two reactive SH groups (SH1 and SH2) in the myosin head heavy chain with N,N'-p-phenylenedimaleimide (pPDM) in the presence of ADP, and by forming a complex with vanadate ion and ADP. At this resolution (approximately 2 nm) the heads of these modified molecules did not appear markedly different from those of the untreated protein, although there was a small increase in the number of straight as opposed to curved heads after cross-linking with pPDM.

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