A new scalable purification procedure for prostatic secretory protein (PSP94) from human seminal plasma.
A simple three-step procedure for the purification of native prostate secretory protein (PSP94) from human seminal plasma is described. The steps are ammonium sulfate fractionation followed by a Macro-Prep S support (cation) flowthrough process and the final Macro-Prep high Q support (anion exchange) chromatography using two step-gradient elution. The benefits of this procedure lie in its simplicity, speed, and relatively inexpensive materials, thus providing advantages over the previously reported schemes. The purity of the product as judged by single band on SDS-polyacrylaminde gel electrophoresis was equivalent to that attained by analytical HPLC anion exchange procedure. Yields were in the range of 18-25 mg highly pure PSP94 per 50 ml of seminal plasma. The desalted, lyophilized, purified PSP94 sample was characterized by SDS-PAGE, Western blot, and N-terminal sequencing. All parameters tested confirm its identity. Preliminary data show that this procedure is suitable for a large-scale production. The direct quantitation of PSP94 by SDS-PAGE and densitometric image analysis at various purification steps for evaluating the recovery of PSP94 is described. The results obtained show that this is an efficient strategy for preparation of highly purified native PSP94.
Baijal-Gupta
M
M
Procyon Biopharma Inc., London, Ontario, Canada.
Fraser
J E
JE
Clarke
M W
MW
Xuan
J W
JW
Finkelman
M A
MA
D015153:Blotting, Western; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004961:Estramustine / Q000737:chemistry*; D006801:Humans; D008297:Male; D011467:Prostate / Q000737:chemistry*; D029584:Prostatic Secretory Proteins; D011506:Proteins / Q000302:isolation & purification*; D012661:Semen / Q000737:chemistry*; D029589:Seminal Plasma Proteins
D029584:Prostatic Secretory Proteins; D011506:Proteins; D029589:Seminal Plasma Proteins; C043076:beta-microseminoprotein; D004961:Estramustine