Serum bound forms of PSP94 (prostate secretory protein of 94 amino acids) in prostate cancer patients.
PSP94 (prostate secretory protein of 94 amino acids) was regarded as a possible prostate cancer marker, however, it has been controversial. All prior studies were designed to test the free form in serum using antibodies to PSP94. Results presented here demonstrate that PSP94 exists in prostate cancer patients in two forms, free and bound, and that the majority is present as serum bound complexes. This result was demonstrated by using both native and SDS-PAGE analyses of serum proteins from prostate cancer patients. Chromatographic separation of serum total proteins by a molecular sieve column generated two peaks (peak I and II), which were reactive with rabbit antiserum to human PSP94 in Western blot experiments. Peak I was eluted before the IgG fraction at a molecular weight larger than 150 kDa, and peak II appeared after serum albumin ( approximately 67 kDa) was eluted. By using a biotinylated PSP94 as an indicator of the free form of PSP94, we demonstrate that peak I contains serum PSP94-bound complexes and peak II is likely the free form of serum PSP94. Since the molecular weight of serum PSP94-bound complexes is close to IgG during molecular sieve separation, serum PSP94 complexes were further purified through two rounds of protein A column separation, followed by DEAE-ion exchange column chromatography. In vitro dissociation tests of the purified PSP94-bound complexes showed that the binding of serum PSP94-complexes is probably via disulfide bonds and is chemically stable. The results presented here indicate that serum PSP94-bound complexes must be considered in evaluating the clinical utility of PSP94 as a prostate cancer marker.
Wu
D
D
Department of Surgery, University of Western Ontario, London, Ontario N6A 4G5, Canada.
Guo
Y
Y
Chambers
A F
AF
Izawa
J I
JI
Chin
J L
JL
Xuan
J W
JW
D000906:Antibodies / Q000097:blood; D014408:Biomarkers, Tumor / Q000097:blood*; D015153:Blotting, Western; D002853:Chromatography, Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008297:Male; D011471:Prostatic Neoplasms / Q000097:blood*; D029584:Prostatic Secretory Proteins; D011506:Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D029589:Seminal Plasma Proteins
D000906:Antibodies; D014408:Biomarkers, Tumor; D029584:Prostatic Secretory Proteins; D011506:Proteins; D029589:Seminal Plasma Proteins; C043076:beta-microseminoprotein