Document Detail


The interaction and photolabeling of myosin subfragment 1 with 3'(2')-O-(4-benzoyl)benzoyladenosine 5'-triphosphate.
MedLine Citation:
PMID:  2959659     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The photoprobe 3'(2')-O-(4-benzoyl)benzoyladenosine 5'-triphosphate (Bz2ATP) was used to characterize the nucleotide-binding site of myosin subfragment 1 (SF1). Improved synthesis and purification of Bz2ATP are reported. 1H NMR and ultraviolet spectroscopy show that Bz2ATP is a 60:40 mixture of the 3'(2')-ribose isomers and that the epsilon M261 is 41,000 M-1 cm-1. Bz2ATP is hydrolyzed by SF1 comparably to ATP in the presence of actin or K+, NH4+, or Mg2+ ions; and the product, Bz2ADP, has a single binding site on SF1 (K'a = 3.0 X 10(5) M-1). [3H]Bz2ATP was photoincorporated into SF1 with concomitant loss of K+-EDTA-ATPase activity. Analysis of photolabeled SF1 showed that the three major tryptic peptides (23, 50, and 20 kDa) of the heavy chain fragment and the alkali light chains were labeled. The presence of ATP during irradiation protected only the 50-kDa peptide, indicating that the other peptides were nonspecifically labeled. If Bz2ATP was first trapped on SF1 by cross-linking the reactive thiols, SH1 and SH2, with p-phenylenedimaleimide, only the 50-kDa tryptic peptide was labeled. These results confirm and extend previous observations that [3H]Bz2ATP trapped on SF1 by cobalt(III) phenanthroline photolabeled the same 50-kDa peptide (Mahmood, R., and Yount, R.G. (1984) J. Biol. Chem. 259, 12956-12959). Thus, the 50-kDa peptide is labeled with or without thiol cross-linking, indicating that the relative position of SH1 and SH2 does not affect the labeling pattern.
Authors:
R Mahmood; C Cremo; K L Nakamaye; R G Yount
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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:  The Journal of biological chemistry     Volume:  262     ISSN:  0021-9258     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  1987 Oct 
Date Detail:
Created Date:  1987-11-23     Completed Date:  1987-11-23     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:  14479-86     Citation Subset:  IM    
Affiliation:
Institute of Biological Chemistry, Washington State University, Pullman 99164-4660.
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MeSH Terms
Descriptor/Qualifier:
Adenosine Triphosphatases / antagonists & inhibitors
Adenosine Triphosphate / analogs & derivatives*,  chemical synthesis,  metabolism
Affinity Labels / metabolism*
Binding Sites
Electrophoresis, Polyacrylamide Gel
Magnetic Resonance Spectroscopy
Myosin Subfragments
Myosins / metabolism*
Peptide Fragments / metabolism*
Photolysis
Spectrophotometry, Ultraviolet
Grant Support
ID/Acronym/Agency:
DK-05195/DK/NIDDK NIH HHS
Chemical
Reg. No./Substance:
0/Affinity Labels; 0/Myosin Subfragments; 0/Peptide Fragments; 56-65-5/Adenosine Triphosphate; 81790-82-1/3'-O-(4-benzoyl)benzoyladenosine 5'-triphosphate; EC 3.6.1.-/Adenosine Triphosphatases; EC 3.6.4.1/Myosins

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


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