Casein kinase II activity and polyamine-stimulated protein phosphorylation of cytosolic and plasma membrane proteins in bovine sperm.
A highly purified preparation of sperm cytosolic protein kinase was obtained by repeated chromatography with phosphocellulose. The preferred substrate of the enzyme was casein and the activity was not stimulated by added Ca2+, calmodulin, or cAMP. With casein as substrate, both ATP and GTP served as phosphate donors and the activity was inhibited by low micromolar heparin and stimulated by low millimolar spermine and spermidine. These properties are characteristic of casein kinase II from other cells. Endogenous protein substrates of the enzyme in sperm cytosolic fractions and in plasma membranes were demonstrated by incubating the preparations with [gamma-32P]GTP, under conditions unfavorable to other protein kinases, and analyzing the products by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. Spermine greatly enhanced the phosphorylation of three (55, 92, and 106 kDa) proteins in both cytosolic and plasma membrane preparations. Our results indicate that polyamines play a role in modulating the phosphorylation state of proteins in sperm and may further regulate sperm function through this mechanism.
Chaudhry
P S
PS
Department of Chemistry, New Mexico State University, Las Cruces 88003.
Casillas
E R
ER
D000818:Animals; D001345:Autoradiography; D047388:Casein Kinases; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology*; D000242:Cyclic AMP / Q000494:pharmacology; D003600:Cytosol / Q000187:drug effects / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D006160:Guanosine Triphosphate / Q000494:pharmacology; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D010766:Phosphorylation; D011073:Polyamines / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000302:isolation & purification* / Q000378:metabolism; D013094:Spermatozoa / Q000187:drug effects / Q000201:enzymology*; D013379:Substrate Specificity
D008565:Membrane Proteins; D011073:Polyamines; D047428:Protein Kinase Inhibitors; D006160:Guanosine Triphosphate; D000242:Cyclic AMP; D011494:Protein Kinases; D047388:Casein Kinases