Isolation and properties of a phosphatidylcholine-specific phospholipase C from bull seminal plasma

J Biol Chem. 1986 Jun 5;261(16):7544-9.


A phospholipase C which hydrolyzes [14C]phosphatidylcholine has been purified 1782-fold from 70% ammonium sulfate extract of bull seminal plasma. Purification steps included acid precipitation, chromatography on DEAE-Sephacel, concanavalin A, octyl-Sepharose 4B and Ultrogel AcA 34. The final step provided homogeneous phospholipase C as determined by polyacrylamide gel electrophoresis. The enzyme comprised two subunits, Mr 69,000 and Mr 55,000, respectively. The enzyme had an optimum at pH 7.2 and pI 5.0. EDTA, Cd2+, Pb2+, Ni2+, Fe2+, and Zn2+ inhibited phospholipase C activity. Km and Vmax on p-nitrophenyl phosphorylcholine and phosphatidylcholine substrates were 20 mM and 17 mumol/min/mg of the purified enzyme and 100 microM and 18 mumol/min/mg of the purified enzyme, respectively. The enzyme appeared to be localized in the acrosome as judged by the binding of anti-phospholipase C to the acrosome. This phospholipase C, unlike other known phospholipases (C), did not hydrolyze [1-14C]phosphatidylinositol. The testicular extract of the guinea pig contained inactive phospholipase C which was activated on incubation with acrosin and trypsin but not chymotrypsin.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Acrosin / pharmacology
  • Animals
  • Carbon Radioisotopes
  • Cattle
  • Edetic Acid / pharmacology
  • Hydrolysis
  • Isoelectric Focusing
  • Kinetics
  • Male
  • Molecular Weight
  • Phosphatidylcholines / metabolism*
  • Phosphatidylethanolamines / metabolism
  • Phosphatidylinositols / metabolism
  • Semen / enzymology*
  • Trypsin / pharmacology
  • Type C Phospholipases / analysis
  • Type C Phospholipases / isolation & purification*


  • Carbon Radioisotopes
  • Phosphatidylcholines
  • Phosphatidylethanolamines
  • Phosphatidylinositols
  • Edetic Acid
  • Type C Phospholipases
  • Acrosin
  • Trypsin