Isolation and characterization of the major gel proteins in human semen, semenogelin I and semenogelin II.
Semenogelin I and semenogelin II constitute the major gel-forming proteins in human semen. The gel proteins were rapidly solubilized and separated from spermatozoa in ejaculates collected at pH 9.7 in buffer containing 4 mol/l urea and dithiothreitol. This protected the semenogelins from proteolytic degradation by prostate-specific antigen, and allowed their isolation by affinity chromatography on heparin-Sepharose. Semenogelins I and II were almost selectively retained and eluted partially separated in 0.25 mol/l NaCl. Further purification was achieved by chromatography on Superose. Approximately 10-20 mg semenogelin I and 2-5 mg semenogelin II were recovered from each sample with a purity exceeding 95% as judged by SDS/PAGE. The molecular mass of semenogelin I (49 958 Da) and the major form of semenogelin II (63 539 Da) measured by mass spectrometry was consistent with the reported cDNA data. The occurrence of a second, larger form of semenogelin II was due to asparagine-linked glycosylation. The amino-termini of the purified proteins were blocked, but digestion with pyroglutamate amino-peptidase enabled the identification of amino-terminal sequences consistent with the reported cDNA data. The amino acid compositions of the purified proteins were also consistent with those derived from cDNA data. The absorption coefficients (280 nm, 1%, 1 cm) for semenogelins I and II were 5.5 and 5.4, respectively, and the isoelectric point was above pH 9.5 for both proteins.
Malm
J
J
Department of Clinical Chemistry, Lund University, University Hospital, Malmö, Sweden.
Hellman
J
J
Magnusson
H
H
Laurell
C B
CB
Lilja
H
H
D000581:Amidohydrolases / Q000378:metabolism; D000596:Amino Acids / Q000032:analysis; D005782:Gels / Q000737:chemistry; D006031:Glycosylation; D012739:Gonadal Steroid Hormones / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D008297:Male; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D017430:Prostate-Specific Antigen / Q000378:metabolism; D011498:Protein Precursors / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D012661:Semen / Q000737:chemistry*; D029589:Seminal Plasma Proteins; D029607:Seminal Vesicle Secretory Proteins; D012995:Solubility
D000596:Amino Acids; D005782:Gels; D012739:Gonadal Steroid Hormones; D011498:Protein Precursors; D029589:Seminal Plasma Proteins; D029607:Seminal Vesicle Secretory Proteins; C424482:seminal vesicle-specific antigen; D017430:Prostate-Specific Antigen; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase