Increased DNA unwinding efficiency of bacteriophage T7 DNA helicase mutant protein 4A'/E348K

M T Washington; S S Patel

doi:10.1074/jbc.273.14.7880

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Increased DNA unwinding efficiency of bacteriophage T7 DNA helicase mutant protein 4A'/E348K

Journal article

Open access

Peer reviewed

Increased DNA unwinding efficiency of bacteriophage T7 DNA helicase mutant protein 4A'/E348K

M T Washington and S S Patel

The Journal of biological chemistry, Vol.273(14), pp.7880-7887

04/03/1998

DOI: 10.1074/jbc.273.14.7880

PMID: 9525882

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Abstract

Bacteriophage T7 4A' protein is a DNA helicase that unwinds DNA in a reaction coupled to dTTP hydrolysis. To understand better its mechanism of DNA unwinding, we characterized a set of 4A' mutant proteins (Washington, M. T., Rosenberg, A. H., Griffin, K., Studier, F. W., and Patel, S. S. (1996) J. Biol. Chem. 271, 26825-26834). We showed here, using single turnover DNA unwinding assays, that the 4A'/E348K mutant protein had the unusual property of unwinding DNA (with a 5-6-fold slower rate) despite a significant defect in its dTTPase activity (a 25-30-fold slower rate). Comparing the DNA unwinding rates to the dTTPase rates, we estimated the DNA unwinding efficiencies of both wild-type (about 1 base pair unwound per dTTP hydrolysis) and mutant (4 to 6 base pairs unwound per dTTP hydrolysis). Thus the mutant had a 4-6-fold improvement in its DNA unwinding efficiency over that of the wild-type. We believe that this mutant undergoes less slippage (uncoupled dTTP hydrolysis) than the wild-type. We speculate that nature has selected for a high rate of DNA unwinding rather than a high efficiency of DNA unwinding. Thus even though the mutant is more efficient at DNA unwinding, the wild-type probably was selected because it unwinds DNA faster.

Hydrolysis

Amino Acid Sequence

DNA Helicases - metabolism

Bacteriophage T7 - genetics

Bacteriophage T7 - metabolism

Base Sequence

DNA, Viral - metabolism

Molecular Sequence Data

Kinetics

Mutation

DNA, Viral - genetics

DNA Helicases - genetics

Details

Title: Subtitle: Increased DNA unwinding efficiency of bacteriophage T7 DNA helicase mutant protein 4A'/E348K
Creators: M T Washington - Department of Biochemistry, The Ohio State University, Columbus, Ohio 43210, USA
S S Patel
Resource Type: Journal article
Publication Details: The Journal of biological chemistry, Vol.273(14), pp.7880-7887
DOI: 10.1074/jbc.273.14.7880
PMID: 9525882
NLM abbreviation: J Biol Chem
ISSN: 0021-9258
eISSN: 1083-351X
Publisher: United States
Grant note: GM55310 / NIGMS NIH HHS
Language: English
Date published: 04/03/1998
Academic Unit: Radiation Oncology; Biochemistry and Molecular Biology
Record Identifier: 9984025298202771

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