THE KINETICS OF TRANSFER OF ESCHERICHIA COLI SINGLE-STRAND DNA BINDING PROTEIN BETWEEN SINGLE-STRANDED DNA MOLECULES.
Item
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Title
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THE KINETICS OF TRANSFER OF ESCHERICHIA COLI SINGLE-STRAND DNA BINDING PROTEIN BETWEEN SINGLE-STRANDED DNA MOLECULES.
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Identifier
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AAI8212213
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identifier
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8212213
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Creator
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SCHNEIDER, ROBERT JAY.
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Contributor
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James G. Wetmur
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Date
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1982
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Language
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English
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Publisher
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City University of New York.
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Subject
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Biology, Microbiology
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Abstract
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The binding of SSB to a ('3)H-labelled 375 nucleotide single-stranded DNA was detected using a nitrocellulose filter binding assay. The binding of protein was sufficiently cooperative that an all-or-none mechanism governs and DNA was either free of or saturated with SSB. The stoichiometry of filter binding agrees with this model. It was found that the first-order kinetics of transfer of protein from donor DNA-protein complexes to recipient DNA of the same size was the same whether using ('3)H-labelled donor or recipient DNA. The rate of transfer of protein from donor to recipient was found to be weakly temperature dependent, independent of salt concentration over a defined range, and inversely proportional to solvent viscosity. These results are consistent with a diffusion-controlled reaction mechanism. When much larger recipient DNA molecules were used, the rate of transfer of SSB was greatly reduced. Also the rate constant for transfer to intact circular single-stranded DNA molecules was not different than that for transfer to linear molecules of about the same length. This indicated that DNA ends are not required for the uptake of SSB. The magnitude and length dependence of the rate constants for protein transfer are incompatible with a mechanism involving uptake of free protein from solution. A model was proposed involving direct transfer of cooperative units of protein from donor to recipient stands.;SSB transfer studies analyzed by electron microscopic visualization were performed with 400 nucleotide single-stranded DNA molecules and circular single-stranded bacteriophage G4 DNA molecules. The transfer of protein from 400 nucleotide DNA donors to G4 recipients and vice-versa are presented in electron micrographs and a statistical compilation of the transfer products. They were found to be consistent with the model presented.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
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Program
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Biomedical Sciences
