Abstract
SspB, a specificity factor for the ATP-dependent ClpXP protease, stimulates proteolysis of protein substrates bearing the ssrA degradation tag. The SspB protein is shown here to form a stable homodimer with two independent binding sites for ssrA-tagged proteins or peptides. SspB by itself binds to ClpX and stimulates the ATPase activity of this enzyme. In the presence of ATPgammaS, a ternary complex of SspB, GFP-ssrA, and the ClpX ATPase was sufficiently stable to isolate by gel-filtration or ion-exchange chromatography. This complex consists of one SspB dimer, two molecules of GFP-ssrA, and one ClpX hexamer. SspB dimers do not commit bound substrates to ClpXP degradation but increase the affinity and cooperativity of binding of ssrA-tagged substrates to ClpX, facilitating enhanced degradation at low substrate concentrations.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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ATPases Associated with Diverse Cellular Activities
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Adenosine Triphosphatases / metabolism*
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Carrier Proteins / metabolism*
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Cleavage And Polyadenylation Specificity Factor / metabolism*
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Dimerization
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Endopeptidase Clp
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Escherichia coli Proteins / metabolism*
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Green Fluorescent Proteins
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Luminescent Proteins
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Molecular Chaperones
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Protein Binding
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Proteins / metabolism
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Serine Endopeptidases / metabolism*
Substances
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Carrier Proteins
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Cleavage And Polyadenylation Specificity Factor
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Escherichia coli Proteins
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Luminescent Proteins
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Molecular Chaperones
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Proteins
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sspB protein, E coli
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Green Fluorescent Proteins
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Serine Endopeptidases
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ClpXP protease, E coli
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Endopeptidase Clp
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Adenosine Triphosphatases
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ClpX protein, E coli
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ATPases Associated with Diverse Cellular Activities