ANALYSIS OF EPITOPE STRUCTURE OF PSP94 (PROSTATE SECRETORY PROTEIN OF94 AMINO-ACIDS) .1. IMMUNODOMINANT AND IMMUNO-RECESSIVE AREA

PSP94 is a potential biomarker for evaluating patients with prostate c arcinoma. We have systematically studied the epitope structure of PSP9 4 by using a polyclonal antibody against human PSP94. Results of pepti de mapping and ELISA tests of dose response to rabbit antiserum agains t human PSP94 protein showed that only the N-terminal peptides (N30 an d M23) are immunoreactive while all the synthetic peptides (C28, C10) located closer to the C-terminus are completely devoid of antigenic ac tivity with the polyclonal antibody. These results were confirmed by a nalysis of reciprocal competitive binding of PSP94 polyclonal antibody by the N-terminal peptides (N30 and M23) v. either recombinant GST-PS P94 fusion protein, purified recombinant PSP94, or natural PSP94 prote in. To further delineate the antigenic activity of the N- and C-termin i, we have also expressed N- and C-terminal half of the whale PSP94 (e ach 47 peptides) using the E. coli GST expression system. The recombin ant N47/C47 peptides were released by thrombin cleavage from the GST f usion protein and characterized by Western blotting experiments. Dose response of the recombinant GST-PSP-N47 and -C47 peptides to PSP94 pol yclonal antibody showed differential binding activities. Competitive b inding of these recombinant N47/C47 proteins against the GST-PSP94 pro tein demonstrates that the polyclonal antibody has a higher affinity f or the N47 peptide than the C47 peptide. Based on the immunological st udies of both synthetic peptides and recombinant PSP94- N/C terminal p roteins, we propose an epitope structure of human PSP94 with an immno- dominant N-terminus and an immune-recessive C-terminus. (C) 1997 Wiley -Liss, Inc.