Abstract
We have cloned and overexpressed a gene encoding a 43-kDa protein corresponding to the N-terminal fragment of the DNA gyrase B subunit. We show that this protein hydrolyzes ATP and binds coumarin drugs. The hydrolysis of ATP shows distinctly non-Michaelis-Menten kinetics and is consistent with a scheme in which the active form of the protein is a dimer, a conclusion supported by molecular weight studies. The coumarin drugs bind very tightly to the 43-kDa fragment, with novobiocin binding to the protein monomer and coumermycin A1 apparently inducing the formation of a dimer. The implications of these results with respect to the mechanism of supercoiling by DNA gyrase and the inhibition of gyrase by coumarin drugs are discussed.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adenosine Triphosphatases / metabolism*
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Adenosine Triphosphate / metabolism*
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Cloning, Molecular
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Coumarins / metabolism*
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DNA Topoisomerases, Type II / genetics
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DNA Topoisomerases, Type II / isolation & purification
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DNA Topoisomerases, Type II / metabolism*
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Electrophoresis, Polyacrylamide Gel
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Escherichia coli / enzymology*
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Kinetics
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Macromolecular Substances
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Mathematics
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Models, Theoretical
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Molecular Weight
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Peptide Fragments / isolation & purification
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Peptide Fragments / metabolism*
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Recombinant Proteins / isolation & purification
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Recombinant Proteins / metabolism
Substances
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Coumarins
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Macromolecular Substances
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Peptide Fragments
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Recombinant Proteins
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Adenosine Triphosphate
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Adenosine Triphosphatases
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DNA Topoisomerases, Type II