Transamidase Reactions Involved in the Enzymic Coagulation of Semen: Isolation of γ-Glutamyl-ε-Lysine Dipeptide from Clotted Secretion Protein of Guinea Pig Seminal Vesicle

Abstract

New supportive evidence is advanced in favor of the hypothesis that the enzymic coagulation of guinea pig semen involves transamidase reactions that result in the formation of 7y-glutamyl-e-lysine intermolecular cross linkages between molecules of a basic protein in seminal vesicle secretion. The dipeptide -y-glutamyl-elysine was isolated in large quantities from proteolytic digests of the coagulated basic vesicular secretion protein that comprises the seminal clot.The seminal vesicle secretions of guinea pigs (1-4), rats (4-9), and mice (10) contain a small group of bulk proteins that are easily separable from one another by chromatographic and electrophoretic procedures. One of these proteins is strongly basic, and can be coagulated by an enzyme from the anterior prostate (coagulating) gland, but not by thrombin (4, 5). This enzymic coagulation process is centrally involved in the postejaculatory clotting of rodent seminal plasma that results in the formation of a vaginal plug when semen is catapulted into the female genital tract at coitus (1,4,5).In 1967, Notides and Williams-Ashman (4) purified the basic coagulable protein from guinea pig vesicular secretion to a state of homogeneity. They observed that, after hydrolysis in strong acid, the amino-acid composition of this basic protein [which is devoid of proline, tyrosine, and cyst(e)ine] was remarkably similar to the amino-acid composition of washed coagulated material formed by the action of the anterior prostate enzyme on a crude mixture of the saline-soluble bulk proteins of seminal vesicle secretion. The native vesicular secretion basic protein has a molecular weight of roughly 17,900, whereas the molecular weight of the coagulated protein is not known. It was observed (4) that clotting of the vesicular secretion basic protein by the anterior prostate enzyme reduced the amide-NH2 from a value of about 21 residues per 17,900 molecular weight to a value of roughly 12 amide-NH2 residues (the number of amide residues represented by glutaminyl or asparaginyl groups was not determined). The enzymically clotted basic protein is not soluble in 8 M urea or in 6 M guanidinium chloride in the absence or presence of thiol compounds at neutral pH (see ref. 1). These facts, taken together, led Notides and Williams-Ashman (4) to postulate that the enzymic clotting of the basic seminal vesicle secretion protein involved the formation of intermolecular linkages similar to those formed in the fibrinoligase (thrombin-activated fibrin stabilizing factor or activated Factor XIII) reaction of mammalian blood clotting (11-13). It is now well established that fibrinoligase is a transamidating enzyme, which catalyzes the production of selective -y-glutamyl-e-lysine crosslinks between fibrin molecules.Some additional new evidence in support of the hypothesis that the guinea pig seminal clot is formed by an enzymic transamidation reaction is outlined below. Unequivocal proof of the formation of peptide cross linkages is provided by the isolation of 'y-glutamyl-e-lysine d...

Heparan Sulfate and Transglutaminase Activity Are Required for the Formation of Covalently Cross-linked Hedgehog Oligomers

“…Indeed, the incomplete inhibition of UD oligomerization by a range of TG inhibitors is not compatible with the mode of de novo direct polymerization of proteins in a singlephased reaction, which constitutes one type of TG-mediated protein cross-linking reaction. Examples for this type of reaction would be the formation of the vaginal plug after copulation in rodents (31) and lobster hemolymph clotting (32). Notably, another type of known TG action is only incompletely affected by inhibitors or analogue substrates of TGs (9) and can be described as the enzymatic "spot-welding" of non-covalent (e.g.…”

Differential alternative splicing of human transglutaminase 4 in benign prostate hyperplasia and prostate cancer

“…A number of studies concerning the nature of postejeculatory plug have recognized that a transglutaminase isoenzyme secreted from prostate gland plays an important role in the reproduction biology of rodents (Williams-Ashman et al, 1972, 1977Williams-Ashman, 1984). Following studies have identified that copulatory plug is formed by transglutaminase 4 (prostate transglutaminase or TGP), a highly glycosylated protein enriched in dorsal prostate of rodents (Seitz et al, 1990).…”

The major clotting protein from guinea pig seminal vesicle contains eight repeats of a 24-amino acid domain.

“…SVP-1 is a clotting protein that serves as the substrate in the formation of the copulatory plug (2,3). Covalent clotting of this protein is catalyzed by a transglutaminase secreted by the anterior prostate (3,4).…”