Isolation, chemical and immunological characterization of two strongly basic nuclear proteins from human spermatozoa

Abstract

Human spermatozoa were solubilized by treatment with urea/guanidine and fractionated by ion-exchange chromatography with Bio-Rex 70 resin. Four fractions were obtained. The acidic proteins pass unretarded, whereas the moderately basic and two strongly basic protein fractions are eluted by means of guanidine gradients. The main protein components of the strongly basic protein fractions have been isolated by gel filtration on Sephadex G-50. The purified proteins have been named human sperm protamine 1 and 2. They contain 47 and 51 aminoacid residues (mol. wt 6280 and 6840), of which 22 and 24 are arginine and 5 and 4 are half cystine residues, respectively. The electrophoretic mobility in urea/polyacrylamide gels is between that of calf thymus histone and salmon protamine. Human protamine 1 and 2 are both auto-antigens as has been detected by a quantitative immunofluorescence inhibition test.

PIP: Chemical and immunological characterization of 2 strongly basic nuclear proteins isolated from human spermatozoa are described. Spermatozoa proteins were solubilized by treatment with urea/ guanidine and fractionated by ion-exchange chromatography with Bio-Rex 70 resin. The initial peak (Fraction 1) was acidic nonabsorbed protein, Fraction 2 was moderately basic, and Fractions 3 and 4 were strongly basic. The acidic proteins passed unretarded whereas the basic proteins were eluted by guanidine gradients. The main protein components of Fractions 3 and 4 were isolated by gel filtration on Sephadex G-50. The purified proteins, named human sperm protamine 1 and 2, contain 47 and 51 aminoacid residues (molecular weight 6277 and 6844) of which 22 and 24 are arginine and 5 and 4 are half cystine residues, respectively. The electrophoretic mobility in urea/polyacrylamide gels is between that of calf thymus histone and salmon protamine. Quantitative immunofluorescence inhibition tests revealed that protamine 1 and 2 are both autoantigens.