Molecular mechanism of epididymal protease inhibitor modulating the liquefaction of human semen.
- Author:
Zeng-Jun WANG
1
;
Wei ZHANG
;
Ning-Han FENG
;
Ning-Hong SONG
;
Hong-Fei WU
;
Yuan-Geng SUI
Author Information
- Publication Type:Journal Article
- MeSH: Animals; Antibodies; pharmacology; Autoradiography; Humans; Hydrolysis; Male; Prostate-Specific Antigen; metabolism; Proteinase Inhibitory Proteins, Secretory; genetics; immunology; metabolism; Rabbits; Recombinant Proteins; genetics; metabolism; Semen; cytology; metabolism; Seminal Vesicle Secretory Proteins; metabolism; Spermatozoa; metabolism
- From: Asian Journal of Andrology 2008;10(5):770-775
- CountryChina
- Language:English
-
Abstract:
AIMTo study the molecular mechanism of epididymal protease inhibitor (Eppin) modulating the process of prostate specific antigen (PSA) digesting semenogelin (Sg).
METHODSHuman Sg cDNA (nucleotides 82-849) and Eppin cDNA (nucleotides 70-723) were generated by polymerase chain reaction (PCR) and cloned into pET-100D/TOPO. Recombinant Eppin and Sg (rEppin and rSg) were produced by BL21 (DE3). The association of Eppin with Sg was studied by far-western immunoblot and radioautography. In vitro the digestion of rSg by PSA in the presence or absence of rEppin was studied. The effect of anti-Q20E (N-terminal) and C-terminal of Eppin on Eppin-Sg binding was monitored.
RESULTSEppin binds Sg on the surface of human spermatozoa with the C-terminal of Eppin (amino acids 75-133). rSg was digested with PSA and many low molecular weight fragments were produced. When rEppin is bound to rSg, then digested by PSA, incomplete digestion and a 15-kDa fragment results. Antibody binding to the N-terminal of rEppin did not affect rSg digestion. Addition of antibodies to the C-terminal of rEppin inhibited the modulating effect of rEppin.
CONCLUSIONEppin protects a 15-kDa fragment of rSg from hydrolysis by PSA.
