The Doc1 subunit is a processivity factor for the anaphase-promoting complex.

Ubiquitin-mediated proteolysis of securin and mitotic cyclins is essential for exit from mitosis. The final step in ubiquitination of these and other proteins is catalysed by the anaphase-promoting complex (APC), a multi-subunit ubiquitin-protein ligase (E3). Little is known about the molecular reaction resulting in APC-dependent substrate ubiquitination or the role ...
of individual APC subunits in the reaction. Using a well-defined in vitro system, we show that highly purified APC from Saccharomyces cerevisiae ubiquitinates a model cyclin substrate in a processive manner. Analysis of mutant APC lacking the Doc1/Apc10 subunit (APC(doc1 Delta)) indicates that Doc1 is required for processivity. The specific molecular defect in APC(doc1 Delta) is identified by a large increase in apparent K(M) for the cyclin substrate relative to the wild-type enzyme. This suggests that Doc1 stimulates processivity by limiting substrate dissociation. Addition of recombinant Doc1 to APC(doc1 Delta) fully restores enzyme function. Doc1-related domains are found in mechanistically distinct ubiquitin-ligase enzymes and may generally stimulate ubiquitination by contributing to substrate-enzyme affinity.
Mesh Terms:
Anaphase, Cell Cycle, Cell Cycle Proteins, Dose-Response Relationship, Drug, Kinetics, Models, Biological, Mutation, Protein Binding, Protein Structure, Tertiary, Recombinant Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Substrate Specificity, Time Factors, Ubiquitin, Ubiquitin-Protein Ligase Complexes, Ubiquitin-Protein Ligases
Nat. Cell Biol.
Date: Nov. 01, 2002
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