A structural model for the prostate disease marker, human prostate‐specific antigen

Abstract

Prostate-specific antigen (PSA) provides an excellent serum marker for prostate cancer, the most frequent form of cancer in American males. PSA is a 237-residue protease based on sequence homology to kallikrein-like enzymes. To predict the 3-dimensional structure of PSA, homology modeling studies were performed based on sequence and structural alignments with tonin, pancreatic kallikrein, chymotrypsin, and trypsin. The structurally conserved regions of the 4 reference X-ray proteins provided the core structure of PSA, whereas the loop structures were modeled on the loops of tonin and kallikrein. The unique "kallikrein loop" insert, between Ser 95b and Pro 95k of kallikrein, was constructed using molecular mechanics, dynamics, and electrostatics calculations. In the resulting PSA structure, the catalytic triad, involving residues His 57, Asp 102, and Ser 195, and hydrophobic and electrostatic interactions typical of serine proteases were extremely well conserved. Similarly, the 5 disulfide bonds of kallikrein were also conserved in PSA. These results, together with the fact that no major steric clashes arose during the modeling process, provide strong evidence for the validity of the PSA model. Calculation of the electrostatic potential contours of kallikrein and PSA was carried out using the finite difference Poisson-Boltzmann method. The calculations revealed matching areas of negative potential near the catalytic triad, but differences in the positive potential surrounding the active site. The PSA glycosylation site, Asn 61, is fully accessible to the solvent and is enclosed in a positive region of the isopotential map. The bottom of the substrate specificity pocket, residue S1, is a serine (Ser 189) as in chymotrypsin, rather than aspartate (Asp 189) as in tonin, kallikrein, and trypsin. This fact, plus other features of the S1 binding-pocket region, suggest that PSA would prefer substrates with hydrophobic residues at the P1 position. The location of a potential zinc ion binding site involving the side chain of histidines 91, 101, and 233 is also suggested. This PSA model should facilitate the understanding and prediction of structural and functional properties of this important cancer marker.

Thiophilic-interaction chromatography of enzymatically active tissue prostate-specific antigen (T-PSA) and its modulation by zinc ions

“…The limited availability of prostate tissue samples, and the difficulty in recovery of sufficient quantities of T-PSA, prevented us from evaluating the effect on enzymatic activity of T-PSA of the full panel of trace metals. Binding of Zn 2+ to the amino acid residues of the catalytic triad of PSA (His 91, His 101 or His 233) could distort the architecture through minor movements of His 101 which is immediately adjacent to the active site residue, Asp 102 [42]. Furthermore, the binding of a Zn 2+ ion in this part of the molecule may modify the conformation of the loop 95, blocking the entrance to the specificity pocket [43].…”

Activation and enzymatic characterization of recombinant human kallikrein 8

“…The predicted Zn 2q -binding sites of hK2 and hK3 are composed of three His residues (Villoutreix et al, 1994;Lovgren et al, 1999), but sequence analysis shows that hK8 has only one His residue close to these corresponding positions ). hK5, 7 and 14 also possess only one His residue.…”

Roles of the P1, P2, and P3 Residues in Determining Inhibitory Specificity of Kallistatin toward Human Tissue Kallikrein

“…The coordinates of the kallikrein loop from a theoretical molecular model of prostate-specific antigen (1pfa.pdb), obtained from the Protein Data Bank, were used as a template to model that of human tissue kallikrein. The method for generating the kallikrein loop of prostate-specific antigen was described in previous studies (33). The final model of tissue kallikrein was then refined by side chain torsion relieving and energy minimization.…”