Activation of latent protease function of Pro-HK2, but not Pro-PSA, involves autoprocessing

BACKGROUND. Human glandular kallikrein 2 (hK2) and prostate-specific antige n (PSA) are members of an extensive kallikrein family of proteases. Both pr oteases are secreted as zymogens or proenzymes containing a seven amino aci d propeptide that must be proteolytically removed for enzymatic activation. The physiological proteases that activate pro-hK2 and pro-PSA are not know n. METHODS. The pro-hK2 peptide sequence is Val-Pro-Leu-Ile-Gln-Ser-Arg (VPLIQ SR). For PSA, the amino acid sequence of the propeptide is Ala-Pro-Leu-Ile- Leu-Ser-Arg (APLILSR). Fluorescent substrates were made by coupling these p eptide sequences to 7-amino-3-methylcoumarin (AMC). The hydrolysis of the V PLIQSR-AMC and APLILSR-AMC substrates by hK2, PSA, and a panel of purified proteases was determined. RESULTS. HK2 readily cleaved the pro-hK2 peptide substrate VPLIQSR-AMC with a rate of hydrolysis that was similar to 8-fold higher than an equimolar a mount of purified trypsin. HK2 also had the highest hydrolysis rate from am ong a group of other trypsin-like proteases. In contrast, neither hK2 nor P SA was able to appreciably cleave the pro-PSA substrate APLILSR-AMC. The pr o-PSA substrate was most readily hydrolyzed by urokinase and trypsin. CONCLUSIONS. HK2 can hydrolyze the pro-hK2 substrate suggesting that matura tion of pro-hK2 to enzymatically active hK2 involves autoprocessing, As exp ected, PSA, a chymotrypsin-like protease, was unable to hydrolyze either of the propeptide substrates. Therefore, it is unlikely that PSA can auto-pro cess its own enzymatic function. HK2 has trypsin-like specificity but was u nable to hydrolyze the pro-PSA substrate. These results raise the possibili ty that an additional processing protease may be required to fully process PSA to an enzymatically active form. (C) 2001 Wiley-Liss, Inc.