GENERATION, PURIFICATION, AND CHARACTERIZATION OF A RECOMBINANT SOURCE OF HUMAN PROSTATE-SPECIFIC ANTIGEN

Human prostate-specific antigen (PSA), a 33- to 34-kDa serine proteina se with extensive homology to glandular kallikrein, is a single-chain glycoprotein that contains 7% carbohydrate. The presence of PSA in the serum of patients with prostatic cancer is widely employed as a marke r of disease status. PSA has also been thought of as a possible target for use in active specific immunotherapy protocols. To date, the sour ce of PSA employed has been seminal fluid from different individuals; this has raised concerns about differences among PSA batches for stand ardization of assays. This report is the first description of the prod uction and the purification of a recombinant source of PSA using a bac ulovirus expression system. A baculovirus recombinant of the cDNA enco ding the full length PSA was expressed in insect cells yielding two ma jor immunoreactive products of 31 and 29 kDa. The latter size conforms to the molecular weight of a core preprotein deduced from the sequenc e of the cDNA insert. The larger protein represents the N-linked glyco sylated form of the preprotein. Western blot analysis showed that both the glycosylated and aglycosylated forms of PSA reacted with a polycl onal and two different monoclonal antibodies specific for PSA, bV-PSA, like commercially available PSA, showed also low-molecular-weight imm unoreactive products when culture supernatants were concentrated or ta ken through steps of purification. bV-PSA was purified to a final prod uct consisting of a major 29-kDa protein and a minor 31-kDa protein. T his recombinant source of PSA will make it possible to further evaluat e the biological, serological, and functional properties of the antige n and may serve as a more standardized source for serum assays to dete ct PSA, bV-PSA may also serve as a potential source for immunogen in a ctive specific immunotherapy protocols. (C) 1995 Wiley-Liss, Inc.