Enzymatic action of prostate-specific antigen (PSA or hK3): substrate specificity and regulation by Zn(2+), a tight-binding inhibitor.
BACKGROUND
In semen, prostate-specific antigen (PSA or hK3) digests the gel proteins semenogelin I and II, resulting in liquefaction and the release of motile spermatozoa. We characterized the substrate specificity and zinc-mediated inhibition of PSA.


METHODS
The proteolysis of human semenogelin I (SgI) and II (SgII) by PSA was characterized by purification of generated SgI and SgII fragments, N-terminal sequencing, and mass spectrometry. Zn(2+)-inhibition of PSA was studied using a chromogenic substrate.


RESULTS
Eighteen cleavage sites in SgI and 16 in SgII were identified. Cleavages were identified mainly as the C-terminal of certain tyrosine and glutamine residues, but also the C-terminal of histidine, aspartic acid, leucine, serine, and asparagine residues. No cleavages were identified at any arginine, lysine, phenylalanine, tryptophan, or methionine residues, indicating that the substrate specificity of PSA is distinct from that of trypsin, chymotrypsin, tissue kallkrein (hK1), and kallikrein 2 (hK2). Zn(2+) ions have a dramatic effect on PSA activity; the data indicate that Zn(2+) is a tight-binding inhibitor of PSA activity.


CONCLUSIONS
The data will enable the optimized design of PSA activity assays, which may prove instrumental to uncovering the role of PSA in cancer and reproduction. The inhibition data indicate that Zn(2+) could regulate PSA activity, which may prove important in the development of efficient inhibitors of PSA activity.
Malm
J
J
Section for Clinical Chemistry, Department of Laboratory Medicine, Lund University, University Hospital Malmö, Malmö, Sweden. johan.malm@klkemi.mas.lu.se
Hellman
J
J
Hogg
P
P
Lilja
H
H
D000595:Amino Acid Sequence; D002413:Cations, Divalent; D004791:Enzyme Inhibitors / Q000378:metabolism*; D004798:Enzymes; D012739:Gonadal Steroid Hormones / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D017430:Prostate-Specific Antigen / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012661:Semen; D029589:Seminal Plasma Proteins; D029607:Seminal Vesicle Secretory Proteins; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D015032:Zinc / Q000378:metabolism*; D000514:alpha 1-Antichymotrypsin / Q000378:metabolism
D002413:Cations, Divalent; D004791:Enzyme Inhibitors; D004798:Enzymes; D012739:Gonadal Steroid Hormones; D029589:Seminal Plasma Proteins; D029607:Seminal Vesicle Secretory Proteins; D000514:alpha 1-Antichymotrypsin; C424482:seminal vesicle-specific antigen; D017430:Prostate-Specific Antigen; D015032:Zinc