Document Detail

Novel configuration of a Myosin-II transient intermediate analogue revealed by quick-freeze replica electron microscopy.
MedLine Citation:
PMID:  23211187     Owner:  NLM     Status:  Publisher    
In this paper, we described our attempt to characterize rough 3-D feature of the structural analogue of the key-intermediate of myosin's crossbridge cycle. Utilizing quick-freeze deep-etch replica electron microscopy, we observed that actin-attached myosin during in vitro sliding was bent superficially as postulated in the conventional hypothesis, but toward the opposite direction of the putative pre-power-stroke configuration like ADP/inorganic-vanadate-bound myosin. We searched for the conformational species with similar appearance by itself and found that SH1-SH2-crosslinked myosin is a good candidate. To characterize such small-sized asymmetric structures, we employed a new pattern-recognition procedure that accommodates the metal-replicated samples. In this method, the best-matched views to the target microscopic images were selected from a comprehensive set of images simulated from known atomic coordinates of relevant proteins. Together with effective morphological-filtering, we could define the conformational species and the view-angles of the catalytic-domain and the lever-arm cropped from averaged images of disulfide-crosslinked myosin. While the catalytic-domain of the new conformer closely resembled pPDM-treated but SH2-Lys705-crosslinked structure (1L20); a minor product of the same crosslinking reaction, the lever-arm projected differently. Utilizing separately determined view-angles of the catalytic-domain and the lever-arm, we built a model of disulfide-crosslinked myosin. Further combination with the "displacement-mapping" procedure enabled us to reconstruct the global 3-D envelope of the unusual structure whose lever-arm orientation is compatible with our reports on actin-sliding crossbridge structure. Assuming this conformer as the structural analogue of the transient intermediate during actin-sliding, the power-stroke of the lever-arm might accompany the reversal of the disorganized SH1-helix.
Yoshitaka Kimori; Norio Baba; Eisaku Katayama
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Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2012-12-5
Journal Detail:
Title:  The Biochemical journal     Volume:  -     ISSN:  1470-8728     ISO Abbreviation:  Biochem. J.     Publication Date:  2012 Dec 
Date Detail:
Created Date:  2012-12-5     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  2984726R     Medline TA:  Biochem J     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
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