Article abstract
Nature Cell Biology 4, 880 - 887 (2002)
Published online: 28 October 2002 | doi:10.1038/ncb871
The Doc1 subunit is a processivity factor for the anaphase-promoting complex
Christopher W. Carroll1 & David O. Morgan1
Abstract
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 (APCdoc1
) indicates that Doc1 is required for processivity. The specific molecular defect in APCdoc1 is identified by a large increase in apparent KM 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 APCdoc1 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.
- Departments of Physiology and Biochemistry & Biophysics, University of California, San Francisco, CA 94143-0444, USA
Correspondence to: David O. Morgan1 e-mail: dmorgan@cgl.ucsf.edu
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