Purification of maturation-promoting factor, an intracellular regulator of early mitotic events.
Maturation-promoting factor causes germinal vesicle breakdown when injected into Xenopus oocytes and can induce metaphase in a cell-free system. The cell-free assay was used to monitor maturation-promoting factor during its purification from unfertilized Xenopus eggs. Ammonium sulfate precipitation and six chromatographic procedures resulted in a preparation purified greater than 3000-fold that could induce germinal vesicle breakdown within 2 hr when injected into cycloheximide-treated oocytes. Proteins of 45 kDa and 32 kDa were correlated with fractions of highest activity in both assays. These fractions contained a protein kinase activity able to phosphorylate the endogenous 45-kDa protein, as well as histone H1, phosphatase inhibitor 1, and casein. The highly purified preparations described here should help to identify the mechanism of action of maturation-promoting factor and to elucidate the role of protein kinases in the induction of metaphase.
Lohka
M J
MJ
Department of Pharmacology, University of Colorado School of Medicine, Denver 80262.
Hayes
M K
MK
Maller
J L
JL
D000818:Animals; D002845:Chromatography; D003513:Cycloheximide / Q000494:pharmacology; D006133:Growth Substances / Q000302:isolation & purification*; D016200:Maturation-Promoting Factor; D008938:Mitosis; D009865:Oocytes / Q000187:drug effects; D014982:Xenopus laevis
D006133:Growth Substances; D003513:Cycloheximide; D016200:Maturation-Promoting Factor