Rabbit testis proacrosin. Purification, molecular weight estimation, and amino acid and carbohydrate composition of the molecule.

Rabbit testis contains a single molecular form of proacrosin which could be extracted at acid pH and purified 145-fold by Sephadex G-100, SE-Sephadex, and concanavalin A-Sepharose affinity column chromatography. The purified proacrosin was judged to be homogeneous on the basis of chromatographic analyses, disc and sodium dodecyl sulfate-polyacrylamide disc gel electrophoresis, and immunodiffusion analysis. The proacrosin preparation was enzymatically inactive, but could be activated either by trypsin activation or autoactivation at pH 8.5. The molecular weight of the purified proacrosin was estimated to be 67,000 to 68,000 by gel chromatography on a calibrated Sephadex G-100 column, polyacrylamide gel electrophoresis in sodium dodecyl sulfate, and sedimentation equilibrium analysis, and was in agreement with the molecular weight calculated by amino acid and carbohydrate content. The apparent molecular weight of acrosin formed by complete autoactivation of proacrosin was estimated to be about 34,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Sephadex G-100 gel chromatography. Those results suggested that the proacrosin may contain two identical polypeptide chains of about M, = 34,000. Rabbit proacrosin was composed of most of the common amino acids, including relatively high proportions of glutamic acid, aspartic acid, serine, leucine, and alanine, and relatively low proportions of methionine and tryptophan. The amino acid and carbohydrate analyses revealed the presence of 514 amino acid and 25 carbohydrate residues/mol in proacrosin. The polypeptide portion represents approximately 92% of the weight of the molecule with the remainder being carbohydrate in nature consisting of mannose, galactose, fucose, glucosamine, and sialic acid.