Purification and characterization of a calcium transport inhibitor protein from bovine seminal plasma.

Abstract

Previous work (Babcock, D. F., Singh, J. P., and Lardy, H. A. (1979) Dev. Biol. 69, 85-93) has shown that bovine seminal fluid contains a component(s) which is capable of preventing or delaying accumulation of extracellular calcium by ejaculated sperm. We report here the component is proteinaceous in nature and has been purified to apparent homogeneity. Conventional purification techniques were employed including ammonium sulfate fractionation, ion-exchange and gel permeation chromatography. The inhibitor protein is a single peptide of Mr = 9,600-10,500 as determined by gel permeation chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Isoelectric focusing in thin layer agarose gels indicates the protein has a pI = 8.3 which is consistent with a determined amino acid composition rich in basic residues. The protein shows no affinity for periodic acid-Schiff reagent and is therefore assumed to contain no carbohydrate. This protein can be distinguished from other previously characterized seminal plasma proteins and it is assumed that this protein is responsible for delaying uptake of calcium into ejaculated sperm by altering a component(s) of the sperm plasma membrane which serves to actively transport calcium into these cells.