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Human glandular kallikrein, hK2, shows arginine‐restricted specificity and forms complexes with plasma protease inhibitors
Author(s) -
Mikolajczyk Stephen D.,
Millar Lisa S.,
Kumar Abhay,
Saedi Mohammad S.
Publication year - 1998
Publication title -
the prostate
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.295
H-Index - 123
eISSN - 1097-0045
pISSN - 0270-4137
DOI - 10.1002/(sici)1097-0045(19980101)34:1<44::aid-pros6>3.0.co;2-k
Subject(s) - kallikrein , protease , human plasma , arginine , endocrinology , biology , medicine , enzyme , chemistry , biochemistry , amino acid , chromatography
BACKGROUND Human glandular kallikrein (hK2) is a new potential marker for prostate cancer. It is a serine protease expressed in human prostate epithelial cells which has 78% sequence identity with prostate‐specific antigen (PSA). PSA is a widely used biochemical marker for prostate cancer. METHODS Recombinant hK2 expressed in mammalian cells was purified to homogeneity by immunoaffinity chromatography, using an anti‐hK2 mAb. hK2 enzymatic specificity was determined on peptide substrates by N‐terminal amino acid sequencing. hK2 complexes were analyzed by SDS‐PAGE and Western blots. RESULTS hK2 was found to cleave peptide substrates exclusively at selected arginine residues. An amidolytic activity of 4,100 pmol/min per μg hK2 was obtained on the chromogenic substrate H‐D‐Pro‐Phe‐Arg‐ p ‐nitroanilide, while no activity was found on methoxysuccinyl‐Arg‐Pro‐Tyr‐ p ‐nitroanilide, a chymotrypsin substrate used to measure PSA activity. hK2 complexed completely with α 1 ‐antichymotrypsin and α 2 ‐antiplasmin after 4 hr at 37°C, but showed no detectable complex with antithrombin III and α 1 ‐protease inhibitor under these conditions. hK2 also formed a rapid complex with α 2 ‐macroglobulin. CONCLUSIONS These results demonstrate that hK2 is an active protease with arginine‐selective specificity, which forms covalent complexes with plasma protease inhibitors. Prostate 34:44–50, 1998. © 1998 Wiley‐Liss, Inc.