Phospholipase C from human sperm specific for phosphoinositides.
Human sperm lysates were incubated in the presence of 1-[14C]stearoyl-2-acyl-sn-glycero-3-phosphocholine, 1-[14C]stearoyl-2-acyl-sn-glycero-3-phosphoethanolamine or 1-[14C]stearoyl-2-acyl-sn-glycero-3-phosphoinositol. Only the latter substrate was hydrolyzed to a significant extent, with a concomitant formation of 1-[14C]stearoyl-2-acyl-sn-glycerol. Furthermore, incubation of phosphatidyl[3H]inositol under the same conditions was accompanied by the formation, in roughly equal amounts, of [3H]inositol 1-phosphate and [3H]inositol 1:2-cyclic monophosphate. Finally [32P]phosphatidylinositol 4-phosphate and [32P]phosphatidylinositol 4,5-bisphosphate were degraded into [32P]inositol 1,4-bisphosphate and [32P]inositol 1,4,5-trisphosphate, respectively. The phosphoinositide-specific phospholipase C was activated by calcium (optimal concentration 5-10 mM) and inhibited by EGTA, although endogenous calcium supported a half-maximal activity. The enzyme displayed an optimal pH of 6.0 and an apparent Km of 0.08 mM. Its specific activity was around 10 nmol/min per mg protein, which is approximately the same as that found in human blood platelets. Subcellular fractionation revealed that 55% of the enzyme was solubilized under conditions where 80% of acrosin appeared in the supernatants. The majority of the particulate phospholipase C activity (37% of total) was found in the 1000 X g pellet, which contained only 8% of total acrosin activity. Further fractionation of spermatozoa into heads and tails indicated no specific enrichment of phospholipase C activity in any of these two fractions. However, owing to a 4-fold higher protein content in the head compared to the tail fraction, it is concluded that about 80% of particulate phospholipase C activity is located in sperm head. The physiological significance of this enzyme is discussed in relation to a possible role in acrosome reaction and (or) in egg fertilization.
D006801:Humans; D006863:Hydrogen-Ion Concentration; D008297:Male; D010713:Phosphatidylcholines / Q000378:metabolism; D010714:Phosphatidylethanolamines / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism*; D013094:Spermatozoa / Q000201:enzymology*; D013347:Subcellular Fractions / Q000201:enzymology; D013379:Substrate Specificity; D010738:Type C Phospholipases / Q000378:metabolism*
D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010716:Phosphatidylinositols; D010738:Type C Phospholipases