Parital characterization of the rat anti-encephalitogenic protein (RSCP)

A protein antigenically similar to the anti-encephalitogenic bovine spinal cord protein (BSCP) was detected in saline extracts of rat nervous tissues by immunodiffusion analyses using a rabbit anti-BSCP serum. Rat SCP (RSCP) appears to be evenly distributed throughout all parts of the rat nervous system and occurs also in the thymus, thyroid, and adrenal glands. Although immunodiffusion analyses indicated that RSCP shares some antigenic sites with BSCP, anti-RSCP sera reacted only with RSCP, indicating that the major immunogenic determinants of the RSCP are peculiar to the rat and differ from the immunogenic determinants of human, monkey, rabbit, guinea pig, or bovine SCP. Immunoelectrophoretic analyses of concentrated pastes of rat brain (RB) or rat spinal cord (RSC) in agar at pH 8.6 revealed that RSCP occurs in two molecular forms having the electrophoretic mobilities of a serum beta-globulin and a serum gamma-globulin, respectively. However, gamma-RSCP is the predominant component of extracts of brain or spinal cord. Gamma-RSCP was isolated from RB and RSC by a procedure which involved: a) extraction with 0.05 M ammonium acetate buffer, pH 4.0; b) batch absorption of impurities on CM-52 cellulose; c) batch absorption of RSCP on SP-Sephadex, pH 3.5; d) elution of RSCP from SP-Sephadex, pH 5.5; and finally, e) gel filtration on Sephadex G-50 superfine. Purified gamma-RSCP formed one band when analyzed by polyacrylamide electrophoresis in acid gels containing 8 M urea. In contrast, two bands were always present when gamma-RSCP from brain or spinal cord were subjected to SDS-polyacrylamide electrophoresis in 15% gels. The larger of the two components of brain gamma-RSCP had a m.w. of 12,400 daltons, whereas the two components of spinal cord gamma-RSCP were smaller. The molecular sizes of brain RSCP and spinal cord RSCP was estimated by gel filtration chromatography to be 12,400 daltons. The amino acid compositions of gamma-RSCP prepared from RB or RSC were similar except that gamma-RSCP from RSC contained twice as much half-cystine and a slightly higher proportion of basic amino acid than gamma-RSCP from RB.