Sperm-specific Protein Research Articles

The biosynthesis of protamine in trout testis. I. Intracellular site of synthesis.

At a late stage of spermatogenesis, protamine, a sperm-specific protein of low molecular weight and high arginine content ([Formula: see text] of the total residues), appears in the nuclei of testis cells in salmonid and, related fish. This newly synthesized small protein progressively replaces the somatic histones in combination with DNA.Two column chromatography techniques (Bio-Gel P-10 and CM-cellulose) which are capable of completely separating protamine from histones and other testis basic proteins have been employed to identify and characterize newly synthesized protamine. When protamine synthesis begins, histone synthesis is decreased and eventually ceases. By pulse labelling testis cell suspensions for different lengths of time and analyzing the amount of 14C-protamme found in the cytoplasm and in the nucleus, the site of protamine synthesis can be shown to be in the cytoplasm. Further, a cell-free, isolated cytoplasmic fraction can incorporate 14C-arginine into whole protamine molecules, while both an isolated nuclear fraction and high-speed supernatant were relatively inactive. Sedimentation analysis of pulse-labelled testis ribosomes indicates that protamine is synthesized on a class of small polysomes, the disomes. Because of its highly basic nature, the release of protamine from the ribosomes and its subsequent transport into the nucleus, which appears to be rapid, may require special mechanisms.

SOME PHYSICOCHEMICAL PROPERTIES OF SALINE-SOLUBLE PROTEINS IN BOVINE SPERMATOZOA

Summary. Bombardment of bovine spermatozoa with glass beads and subsequent extraction in phosphate buffered sodium chloride yielded soluble sperm-specific proteins (approximately 10% yield) sufficient for physicochemical analysis. Moving boundary electrophoresis revealed three sperm-specific components with mobilities of 2·0, 3·8 and 5·1×10-5 cm2 volt-1 sec-1 at pH 8·6 in barbital (μ = 0·10). The proportions of these components were 20, 50 and 30%, respectively. Sedimentation velocity ultracentrifugal analysis revealed at least three sedimentation gradients. The two major gradients had S20 values of 1·7 and 12·6. The third exhibited polydispersion. Diffusion coefficients of 4·2 and 10·2×10-7 cm2 sec-1 were calculated for two sperm-specific antigens by an agar-gel-diffusion method. Three protein fractions were isolated by chromatographic elution from diethylaminoethyl (deae) cellulose with an ionic gradient. The first protein fraction eluted was not bound to the deae cellulose. It migrated toward the cathode in agar-gel electrophoresis at pH 8·4, and was firmly bound to carboxymethylcellulose at pH 6·0, 7·6 and 11·0. A second protein fraction from the deae cellulose appeared just after the ionic gradient commenced, and a third protein fraction appeared when the ionic gradient approached 0·2 m-sodium chloride.