Modulation of recombinant human prostate-specific antigen: Activation by Hofmeister salts and inhibition by azapeptides

Prostate specific antigen (PSA, also known as human kallikrein 3) is an imp ortant diagnostic indicator of prostatic disease. PSA exhibits low protease activity (> 10(4)-fold less than chymotrypsin) under the usual in vitro as say conditions. In addition, PSA does not react readily with prototypical s erine protease inactivators. We expressed human PSA (rh-PSA) in Escherichia coli and have demonstrated that rh-PSA has properties similar to those of native PSA isolated from human seminal fluid. Both PSA and rh-PSA are > 10( 3)-fold more active in the presence of 1.3 M Na2SO4. This activation is ani on-dependent, following the Hofmeister series when normality is considered: SO42- approximate to citrate > Ac- > Cl- > Br- > I-. The nature of the cat ion has little effect on salt activation. The rate of inactivation of rh-PS A by DFP is 30-fold faster in the presence of 0.9 M Na2SO4, and the rate of inactivation by Suc-Ala-Ala-Pro-Phe-CK is > 20-fold faster under these con ditions. Azapeptides containing Phe or Tyr at position P, also inactivate r h-PSA in the presence of high salt concentrations. These compounds represen t the first described inhibitors designed to utilize the substrate binding subsites of PSA. CD spectroscopy demonstrates that the conformation of rh-P SA changes in the presence of high salt concentrations. Analytical ultracen tifugation and dynamic light scattering indicate that PSA remains monomeric under high-salt conditions. Interestingly, human prostatic fluid contains as much as 150 mu mol citrate/g wet weight, which suggests that salt concen trations may regulate PSA activity in vivo.