Expression, purification, and characterization of deglycosylated human pro-prostate-specific antigen

Wild-type and deglycosylated forms of human prostate-specific antigen were expressed in Chinese hamster ovary (CHO) cells as zymogens. ProPSA was coll ected from conditioned medium and purified using a single cation-exchange c hromatographic step for the deglycosylated form and cation-exchange followe d by gel filtration chromatography for the wild-type form. Recombinant wild -type proPSA produced in CHO cells has an average MW of 34.5 kDa, whereas t he deglycosylated proPSA has a MW of 32.4 kDa. Both forms of proPSA were ac tivated in vitro and the kinetic properties measured for the deglycosylated PSA are very similar to those of the wild-type recombinant PSA and the nat ive PSA isolated from seminal fluid. These results suggest that deglycosyla ted PSA is likely to be very similar to native PSA with respect to its thre e-dimensional structure and will provide a homogeneous protein preparation necessary for X-ray crystallographic analysis, (C) 2000 Academic Press.