Document Detail


ATPase activities and actin-binding properties of subfragments of Acanthamoeba myosin IA.
MedLine Citation:
PMID:  2946692     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Previous studies had led to the conclusion that the globular, single-headed myosins IA and IB from Acanthamoeba castellanii contain two actin-binding sites: one associated with the catalytic site and whose binding to F-actin activates the Mg2+-ATPase activity and a second site whose binding results in the cross-linking of actin filaments and makes the actin-activated ATPase activity positively cooperative with respect to myosin I concentration. We have now prepared a 100,000-Da NH2-terminal peptide and a 30,000-Da COOH-terminal peptide by alpha-chymotryptic digestion of the myosin IA heavy chain. The intact 17,000-Da light chain remained associated with the 100,000-Da fragment, which also contained the serine residue that must be phosphorylated for expression of actin-activated ATPase activity by native myosin IA. The 30,000-Da peptide, which contained 34% glycine and 21% proline, bound to F-actin with a KD less than 0.5 microM in the presence or absence of ATP but had no ATPase activity. The 100,000-Da peptide bound to F-actin with KD = 0.4-0.8 microM in the presence of 2 mM MgATP and KD less than 0.01 microM in the absence of MgATP. In contrast to native myosin IA, neither peptide cross-linked actin filaments. The phosphorylated 100,000-Da peptide had actin-activated ATPase activity with the same Vmax as that of native phosphorylated myosin IA but this activity displayed simple, noncooperative hyperbolic dependence on the actin concentration in contrast to the complex cooperative kinetics observed with native myosin IA. These results provide direct experimental evidence for the presence of two actin-binding sites on myosin IA, as was suggested by enzyme kinetic and filament cross-linking data, and also for the previously proposed mechanism by which monomeric myosins I could support contractile activities.
Authors:
T J Lynch; J P Albanesi; E D Korn; E A Robinson; B Bowers; H Fujisaki
Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  261     ISSN:  0021-9258     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  1986 Dec 
Date Detail:
Created Date:  1987-01-22     Completed Date:  1987-01-22     Revised Date:  2009-11-19    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  17156-62     Citation Subset:  IM    
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MeSH Terms
Descriptor/Qualifier:
Actins / metabolism*
Amino Acids / analysis
Amoeba / metabolism*
Animals
Ca(2+) Mg(2+)-ATPase / metabolism*
Chymotrypsin
Kinetics
Myosin Subfragments
Myosins / isolation & purification,  metabolism*
Peptide Fragments / isolation & purification,  metabolism*
Protein Binding
Chemical
Reg. No./Substance:
0/Actins; 0/Amino Acids; 0/Myosin Subfragments; 0/Peptide Fragments; EC 3.4.21.1/Chymotrypsin; EC 3.6.1.-/Ca(2+) Mg(2+)-ATPase; EC 3.6.4.1/Myosins

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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