Document Detail


Changing the enzymatic activity of T7 endonuclease by mutations at the beta-bridge site: alteration of substrate specificity profile and metal ion requirements by mutation distant from the catalytic domain.
MedLine Citation:
PMID:  15065875     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Phage-encoded resolvase T7 endonuclease I is a structure-specific endonuclease. The enzyme acts on a broad spectrum of substrates with a variety of DNA structures. The enzyme is a dimer with two separated catalytic domains connected by an elongated beta-sheet bridge. The activities of enzymes with mutations in the beta-bridge segment were studied. Mutations that did not affect catalytic domain folding and function but did alter the relative positions of these domains retained catalytic activity but with altered specificity and metal ion dependence. Our results suggest that the enzyme recognizes its substrates by DNA conformation exclusion and offer a simple explanation for the broad substrate specificity of phage resolvase.
Authors:
Chudi Guan; Sanjay Kumar; Rebecca Kucera; Amy Ewel
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Biochemistry     Volume:  43     ISSN:  0006-2960     ISO Abbreviation:  Biochemistry     Publication Date:  2004 Apr 
Date Detail:
Created Date:  2004-04-06     Completed Date:  2004-08-06     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  0370623     Medline TA:  Biochemistry     Country:  United States    
Other Details:
Languages:  eng     Pagination:  4313-22     Citation Subset:  IM    
Affiliation:
New England Biolabs, Inc., 32 Tozer Rd, Beverly, Massachusetts 01915, USA. Guan@neb.com
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MeSH Terms
Descriptor/Qualifier:
Bacteriophage T7 / enzymology*,  genetics*
Base Pair Mismatch / genetics
Binding Sites / genetics
Catalytic Domain / genetics*
DNA, Viral / genetics,  metabolism
Deoxyribonuclease I / biosynthesis,  genetics*,  metabolism*
Enzyme Activation / genetics
Gene Expression Regulation, Viral
Hydrolysis
Magnesium / chemistry*
Manganese / chemistry*
Models, Chemical
Mutagenesis, Site-Directed*
Nucleic Acid Conformation
Nucleic Acid Heteroduplexes / genetics,  metabolism
Substrate Specificity / genetics
Chemical
Reg. No./Substance:
0/DNA, Viral; 0/Nucleic Acid Heteroduplexes; 7439-95-4/Magnesium; 7439-96-5/Manganese; EC 3.1.21.1/Deoxyribonuclease I

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


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