Abstract
Bacteriophage T7 DNA polymerase (gene 5 protein, gp5) interacts with its processivity factor, Escherichia coli thioredoxin, via a unique loop at the tip of the thumb subdomain. We find that this thioredoxin-binding domain is also the site of interaction of the phage-encoded helicase/primase (gp4) and ssDNA binding protein (gp2.5). Thioredoxin itself interacts only weakly with gp4 and gp2.5 but drastically enhances their binding to gp5. The acidic C termini of gp4 and gp2.5 are critical for this interaction in the absence of DNA. However, the C-terminal tail of gp4 is not required for binding to gp5 when the latter is bound to a primer/template. We propose that the thioredoxin-binding domain is a molecular switch that regulates the interaction of T7 DNA polymerase with other proteins of the replisome.
Publication types
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Research Support, U.S. Gov't, Non-P.H.S.
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Bacteriophage T7 / enzymology
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Bacteriophage T7 / genetics
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Base Sequence
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Binding Sites / genetics
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DNA Primase / chemistry*
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DNA Primase / genetics
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DNA Primase / metabolism*
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DNA, Viral / genetics
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DNA-Binding Proteins / chemistry*
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism*
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DNA-Directed DNA Polymerase / chemistry*
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DNA-Directed DNA Polymerase / genetics
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DNA-Directed DNA Polymerase / metabolism*
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Escherichia coli Proteins / chemistry
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Escherichia coli Proteins / metabolism
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Genes, Viral
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Models, Biological
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Models, Molecular
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Mutagenesis
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Protein Conformation
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Sequence Deletion
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Sulfinic Acids
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Thioredoxins / chemistry
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Thioredoxins / metabolism
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Viral Proteins / chemistry*
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Viral Proteins / genetics
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Viral Proteins / metabolism*
Substances
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DNA, Viral
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DNA-Binding Proteins
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Escherichia coli Proteins
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Sulfinic Acids
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Viral Proteins
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gene 2.5 protein, Enterobacteria phage T7
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thiosulfinic acids
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Thioredoxins
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DNA Primase
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bacteriophage T7 induced DNA polymerase
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DNA-Directed DNA Polymerase