Document Detail


Identification of two segments, separated by approximately 45 kilodaltons, of the myosin subfragment 1 heavy chain that can be cross-linked to the SH-1 thiol.
MedLine Citation:
PMID:  3311149     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The thiol-specific photoactivatable reagent 4-(2-iodoacetamido)benzophenone (BPIA) can be selectively incorporated into the SH-1 of myosin subfragment 1 (S1), and upon photolysis an intramolecular cross-link is formed between SH-1 and the N-terminal 25-kDa region of S1. If a Mg2+-nucleotide is present during photolysis, cross-links can be formed either with the 25-kDa or with the central 50-kDa region [Lu, R. C., Moo, L., & Wong, A. G. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 6392-6396]. Heavy chains with these two types of intramolecular cross-links and un-cross-linked heavy chain have different mobility on sodium dodecyl sulfate (NaDodSO4)-polyacrylamide gels and therefore can be purified electrophoretically. Each type of heavy chain was cleaved with Staphylococcus aureus protease, chymotrypsin, or lysyl endopeptidase. The cleavage points were determined on the basis of the molecular weights of weights of peptides containing the N-terminus, which was identified with the use of an antibody. Locations of the cross-links were deduced by comparing the peptide maps of cross-linked and un-cross-linked heavy chains. The results indicate that the segment located about 12-16 kDa from the N-terminus of the heavy chain can be cross-linked to SH-1 via BPIA independently of the presence of a nucleotide, whereas the segment located 57-60 kDa from the N-terminus can be cross-linked to SH-1 only in the presence of a Mg2+-nucleotide. With use of the avidin-biotin system, it has been shown that SH-1 is located 13 nm from the head/rod junction [Sutoh, K., Yamamoto, K., & Wakabayashi, T. (1984) J. Mol. Biol. 178, 323-339]. Since BPIA spans less than 1 nm, our results show that two regions, separated by approximately 400 amino acid residues and located in the 25- and 50-kDa domains of S1, respectively, are also part of the head structure about 12-14 nm from the head/rod junction.
Authors:
K Sutoh; R C Lu
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Biochemistry     Volume:  26     ISSN:  0006-2960     ISO Abbreviation:  Biochemistry     Publication Date:  1987 Jul 
Date Detail:
Created Date:  1987-12-07     Completed Date:  1987-12-07     Revised Date:  2009-11-19    
Medline Journal Info:
Nlm Unique ID:  0370623     Medline TA:  Biochemistry     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  4511-6     Citation Subset:  IM    
Affiliation:
Department of Biophysics and Biochemistry, Faculty of Science, University of Tokyo, Japan.
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MeSH Terms
Descriptor/Qualifier:
Animals
Benzophenones / metabolism*
Cross-Linking Reagents / metabolism*
Molecular Weight
Muscles / metabolism
Myosin Subfragments
Myosins / metabolism*
Peptide Fragments / metabolism*
Peptide Hydrolases
Photochemistry
Protein Conformation
Rabbits
Sulfhydryl Compounds / metabolism
Grant Support
ID/Acronym/Agency:
AM28401/AM/NIADDK NIH HHS
Chemical
Reg. No./Substance:
0/Benzophenones; 0/Cross-Linking Reagents; 0/Myosin Subfragments; 0/Peptide Fragments; 0/Sulfhydryl Compounds; 76809-63-7/benzophenone-4-iodoacetamide; EC 3.4.-/Peptide Hydrolases; EC 3.6.4.1/Myosins

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