Affinity purification of mitotic anaphase-promoting complex/cyclosome on p13Suc1.
A procedure is described for the affinity purification of the mitotic form of anaphase-promoting complex/cyclosome (APC/C) from HeLa cells. It is based on the binding of mitotically phosphorylated APC/C to the phosphate-binding site of p13(suc1), followed by specific elution with a phosphate-containing compound. The procedure is rapid, simple, and yields 50- to 70-fold purification of soluble APC/C, with a approximately 30% recovery of activity.
Hershko
Avram
A
Unit of Biochemistry, Technion-Israel Institute of Technology, Haifa 31096, Israel.
D064173:Anaphase-Promoting Complex-Cyclosome; D018797:Cell Cycle Proteins / Q000302:isolation & purification*; D002846:Chromatography, Affinity / Q000379:methods*; D016213:Cyclins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis / Q000502:physiology*; D010766:Phosphorylation; D029702:Schizosaccharomyces pombe Proteins / Q000302:isolation & purification*; D025801:Ubiquitin / Q000378:metabolism; D043743:Ubiquitin-Protein Ligase Complexes / Q000032:analysis / Q000302:isolation & purification*
D018797:Cell Cycle Proteins; D016213:Cyclins; D029702:Schizosaccharomyces pombe Proteins; C051655:Suc1 protein, S pombe; D025801:Ubiquitin; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome