Document Detail

Sin resolvase catalytic activity and oligomerization state are tightly coupled.
MedLine Citation:
PMID:  20868695     Owner:  NLM     Status:  MEDLINE    
Serine recombinases promote specific DNA rearrangements by a cut-and-paste mechanism that involves cleavage of all four DNA strands at two sites recognized by the enzyme. Dissecting the order and timing of these cleavage events and the steps leading up to them is difficult because the cleavage reaction is readily reversible. Here, we describe assays using activated Sin mutants and a DNA substrate with a 3'-bridging phosphorothiolate modification that renders Sin-mediated DNA cleavage irreversible. We find that activating Sin mutations promote DNA cleavage rather than simply stabilize the cleavage product. Cleavage events at the scissile phosphates on complementary strands of the duplex are tightly coupled, and the overall DNA cleavage rate is strongly dependent on Sin concentration. When combined with analytical ultracentrifugation data, these results suggest that Sin catalytic activity and oligomerization state are tightly linked, and that activating mutations promote formation of a cleavage-competent oligomeric state that is normally formed only transiently within the full synaptic complex.
Kent W Mouw; Andrew M Steiner; Rodolfo Ghirlando; Nan-Sheng Li; Sally-J Rowland; Martin R Boocock; W Marshall Stark; Joseph A Piccirilli; Phoebe A Rice
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't     Date:  2010-09-22
Journal Detail:
Title:  Journal of molecular biology     Volume:  404     ISSN:  1089-8638     ISO Abbreviation:  J. Mol. Biol.     Publication Date:  2010 Nov 
Date Detail:
Created Date:  2010-11-03     Completed Date:  2010-12-06     Revised Date:  2014-09-17    
Medline Journal Info:
Nlm Unique ID:  2985088R     Medline TA:  J Mol Biol     Country:  England    
Other Details:
Languages:  eng     Pagination:  16-33     Citation Subset:  IM    
Copyright Information:
Copyright © 2010 Elsevier Ltd. All rights reserved.
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MeSH Terms
Bacterial Proteins / genetics,  metabolism*
DNA / chemical synthesis,  metabolism*
DNA Nucleotidyltransferases / genetics,  metabolism*
Models, Molecular
Mutant Proteins / genetics,  metabolism
Oligonucleotides / chemical synthesis,  metabolism
Protein Multimerization*
Protein Structure, Quaternary
Recombination, Genetic*
Grant Support
Reg. No./Substance:
0/Bacterial Proteins; 0/Mutant Proteins; 0/Oligonucleotides; 9007-49-2/DNA; EC 2.7.7.-/DNA Nucleotidyltransferases; EC 2.7.7.-/sin protein, Staphylococcus aureus

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