Experimental allergic orchitis: the isolation and partial characterization of an aspermatogenic polypeptide (AP3) with an apparent sequential disease-inducing determinant(s).
An aspermatogenic polypeptide (AP3) capable of inducing experimental allergic orchitis (EAO) in the guinea pig (GP) was purified from GP testes by sequential delipidation, acid extraction, pH precipitation, ammonium sulfate fractionation, trichloroacetic acid precipitation, gel filtration on Sephadex G-75, preparative isoelectric focusing from pH 3-10 followed by isoelectric focusing from pH 7-10, gel filtration on Sephadex G-75 Superfine under reducing conditions, and reduced acid urea gel electrophoresis. Approximately 250 micrograms (BSA equivalents) of AP3 were obtained from 500 g wet weight of GP testes. On 15% reduced acid urea polyacrylamide gels, AP3 appeared as a single band with an Rf of 0.19. SDS-PAGE showed a single band with a mobility corresponding to a m.w. of 12,500 +/- 1500. The isoelectric point, determined during purification, was 9.90 +/- 0.50. Amino acid analysis of AP3 indicates it is a protein. Gas liquid chromatographic analysis failed to reveal the presence of either hexose or hexosamine, indicating that AP3 is probably not a glycopeptide. Two to 5.0 micrograms (BSA equivalents) of AP3 are capable of inducing severe EAO in 100% of GP tested; 1 to 2.0 micrograms (BSA equivalents) induced EAO in 60% of GP tested. Because AP3 appears to be nonglycosylated and the aspermatogenic activity of AP3 is highly resistant to various denaturing conditions including reduction and alkylation, the primary sequence of the polypeptide rather than higher ordered structure may be more important in defining the determinant(s) responsible for its aspermatogenic activity.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D000941:Antigens / Q000302:isolation & purification*; D001324:Autoantigens / Q000008:administration & dosage / Q000302:isolation & purification*; D004195:Disease Models, Animal; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000302:isolation & purification*; D006168:Guinea Pigs; D007525:Isoelectric Focusing; D008297:Male; D009845:Oligospermia / Q000209:etiology / Q000473:pathology; D009920:Orchitis / Q000209:etiology / Q000276:immunology* / Q000473:pathology; D013737:Testis / Q000276:immunology / Q000473:pathology
D000596:Amino Acids; D000941:Antigens; D001324:Autoantigens; D000939:Epitopes