Purification of the mitotic checkpoint complex, an inhibitor of the APC/C from HeLa cells.
The mitotic checkpoint prevents cells with unaligned chromosomes from prematurely exiting mitosis by inhibiting the anaphase-promoting complex/cyclosome (APC/C) from targeting key proteins for ubiquitin-mediated proteolysis. Great efforts have been made recently to identify and characterize the factors involved in this checkpoint pathway. These studies will ultimately lead to a better understanding of cell cycle defects and chromosome instability. We report here a protocol for purification of an inhibitor of the APC/C from HeLa cells, called mitotic checkpoint complex (MCC). Our procedure is based on biochemical purification and characterization of the APC/C inhibitory activity from extracts of HeLa cells.
Sudakin
Valery
V
Institute for Cancer Research, The Fox Chase Cancer Center, Philadelphia, PA, USA.
Yen
Tim J
TJ
D064173:Anaphase-Promoting Complex-Cyclosome; D000970:Antineoplastic Agents / Q000494:pharmacology; D002352:Carrier Proteins / Q000302:isolation & purification* / Q000378:metabolism; D018797:Cell Cycle Proteins; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D006367:HeLa Cells / Q000737:chemistry; D006801:Humans; D008938:Mitosis / Q000502:physiology*; D015739:Nocodazole / Q000494:pharmacology; D009687:Nuclear Proteins; D008941:Spindle Apparatus / Q000378:metabolism; D043743:Ubiquitin-Protein Ligase Complexes / Q000037:antagonists & inhibitors*; D044767:Ubiquitin-Protein Ligases / Q000037:antagonists & inhibitors*
D000970:Antineoplastic Agents; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D009687:Nuclear Proteins; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D044767:Ubiquitin-Protein Ligases; D015739:Nocodazole