title: Purification and characterization of the active precursor of a human sperm motility inhibitor secreted by the seminal vesicles: identity with semenogelin.
abstract: Human seminal plasma contains a sperm motility inhibitor that originates from seminal vesicles as a precursor form. This precursor is degraded into smaller peptides by prostatic proteases shortly after ejaculation. The seminal plasma sperm motility inhibitor (SPMI) precursor was purified by a combination of cation-exchange chromatography on S-Sepharose followed by C4 reverse-phase high-performance liquid chromatography directly from seminal vesicle fluid or washed seminal coagulum. The purification procedure yielded a protein of apparent homogeneity, with a molecular mass of 52 kDa by SDS-PAGE. It migrated as a 105-kDa protein by molecular sieving under denaturing conditions. The purified SPMI precursor was digested by the prostatic protease prostate-specific antigen (PSA), causing a 76 +/- 4% drop in biological activity and transformation into low molecular mass SPMI polypeptides (5-20 kDa) similar to those observed in liquefied semen. The N-terminal amino acid sequences of three degradation peptides were obtained by Edman degradation and found to correspond to residues 45-50, 85-90, and 137-143 of semenogelin, a protein characterized as the major structural component of semen coagulum. The amino acid composition of SPMI precursor was found to be almost identical to that of semenogelin. Moreover, the mass of the precursor was estimated at 49,620 daltons by electrospray-ionization mass spectrometry, a value in close agreement with the expected mass of semenogelin according to its cDNA sequence. The SPMI precursor was found to inhibit sperm motility in a dose-dependent manner, with complete immobilization at 500 U/ml of SPMI. The motility of completely immobilized spermatozoa was partially recovered after washing of the cells. The results suggest that SPMI precursor is the major component of the seminal vesicle secretions and seminal coagulum. It can be degraded by PSA in a manner reminiscent of its processing in whole semen. Taken together these results indicate that the SPMI precursor is semenogelin and that intact semenogelin can immobilize spermatozoa.
authors.affiliation: Urology Research Laboratory, Royal Victoria Hospital, Montréal, Québec, Canada.
authors.forename: M
authors.initials: M
authors.lastname: Robert
authors.forename: C
authors.initials: C
authors.lastname: Gagnon
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D002847:Chromatography, Agarose; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D012739:Gonadal Steroid Hormones / Q000737:chemistry*; D006801:Humans; D015151:Immunoblotting; D008297:Male; D017430:Prostate-Specific Antigen / Q000494:pharmacology; D011498:Protein Precursors / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D029607:Seminal Vesicle Secretory Proteins; D012669:Seminal Vesicles / Q000737:chemistry / Q000378:metabolism*; D017421:Sequence Analysis; D013056:Spectrophotometry, Ultraviolet; D013081:Sperm Motility / Q000502:physiology
chemical_list: D000596:Amino Acids; D004791:Enzyme Inhibitors; D012739:Gonadal Steroid Hormones; D011498:Protein Precursors; D011506:Proteins; D029607:Seminal Vesicle Secretory Proteins; C424482:seminal vesicle-specific antigen; D017430:Prostate-Specific Antigen