The proteins of epididymal luminal fluid and of spermatozoa recovered from different regions of the rat epididymis were examined by polyacrylamide gel electrophoresis under non-denaturing and denaturing conditions. Albumin (A) and four major pre-albumin bands (B-E) were observed in epididymal fluid from the cauda on non-denaturing gels. By comparing the migration of these bands with that of standard globular proteins on denaturing gels, the molecular weight of Bands B and C was estimated to be 16 000, Band D was 30 000 and Band E was 32 000. Bands D and E were apparently glycoproteins since they stained with periodic acid-Schiff's reagent and were bound by an affinity column of Concanavalin A. The pre-albumin proteins (B-E) were of epididymal origin since they (a) were not detected in blood serum, (b) were not detected in testicular extracts and (c) were still found after ligation of the efferent ducts. From the incorporation of radioactive methionine, Bands B and C were shown to be synthesized in the initial segment and caput. The regional distribution of luminal proteins indicated that protein D was added in the caput and cauda and protein E in the cauda. This regional origin of luminal proteins was confirmed by the altered protein profiles consequent upon the reduced fluid flow through the epididymis brought about by ligation of the efferent ducts. The androgen-dependence of epididymal protein synthesis was also investigated using radioactive methionine. Castration had little effect on total protein synthesis but resulted in the specific reduction of the synthesis of proteins B and C. Several changes were observed in the relative amounts of specific proteins extracted from spermatozoa from different regions of the epididymis and several of these proteins had molecular weights identical with those in luminal fluid. However, there was no evidence for any substantial binding to spermatozoa of the pre-albumin proteins (B-E) of luminal fluid.
Frailty in vascular surgery patients predicts a multiplicity of poorer outcomes. Optimal management should include identification of at-risk patients and treatment of modifiable risk factors.
cDNA clones coding for two closely related androgen-dependent sperm-coating glycoproteins secreted by the rat epididymis were selected by screening an epididymal cDNA library constructed in Agt 11 with affinity-purified antibody directed against the glycoproteins. The largest clone of 956 nucleotides provided coding information for a protein of 246 amino acids of which the first 19 residues comprise a putative signal peptide sequence which when cleaved would produce a mature protein of 227 residues and a molecular mass of 26 kDa. Confirmation of the identity of the clone was provided by a match between the amino acid sequence predicted from the cDNA sequence and the actual amino acid sequence determined for a tryptic peptide fragment of one of the pure glycoproteins. It is probable that the primary amino acid sequence of the two glycoproteins is identical. Northern blot and slot-blot analysis revealed that the mRNA for the glycoproteins is approximately 1250 nucleotides long and that the concentration of the mRNA in the epididymis is androgen-dependent. The glycoproteins and their mRNAs were unique to the epididymis as determined by Western and Northern blots, respectively, since signals were absent from skin, brain, liver, kidney, heart, skeletal muscle and testis. Cross-reacting proteins of slightly smaller apparent molecular mass were detected in extracts of mouse and guinea-pig epididymis, but not rabbit or bull epididymis. Comparison with existing protein data bases revealed that the epididymal glycoproteins display significant sequence homology with yeast carboxypeptidase Y.The mammalian epididymis is a single long convoluted tubule which conveys spermatozoa from the testis to the ductus deferens. During passage through this tubule spermatozoa undergo functional maturation which is manifest in the progressive acquisition by the cells of the ability to swim [l, 21, to bind to the zona pellucida of the oocyte [3, 41, and to effect fertilization [5]. A number of studies indicate that functional maturation is not an intrinsic property of the sperm, but is brought about by an interaction of sperm with a variety of androgen-dependent proteins which are secreted by the epididymal epithelium and then become associated with the sperm surface [6-131. We have recently reported the cDNA cloning and primary amino acid sequence of two androgen-dependent epididymal secretory proteins of 18.5 kDa which are thought to be involved in this way in sperm development [ 141. However, particular attention has also been focused on some acidic glycoproteins which have been given several names such as proteins D and E [15, 161, acidic epididymal glycoprotein [7], protein IV [17], sialoprotein [18], and 32 K protein [19]. The glycoproteins have been variously reported to have isoelectric points between 4.0 and 5.1 and molecular masses in the range of 27-37 kDa
Carnitine and glycerylphosphorylcholine (GPC) levels have been measured in the reproductive tissues of male rats, with special reference to the epididymis. The increase in concentration of these compounds in reproductive tissues during the prepubertal and pubertal periods has been examined and their distribution along the epididymis has been determined in mature animals. Carnitine accumulates in the epididymal plasma in the cauda epididymidis whilst GPC accumulates in the caput epididymidis. By developing a new technique for the collection and separation of epididymal spermatozoa and undiluted fluid from the cauda epididymidis, the concentrations of carnitine, acetylcarnitine and GPC have been measured and the free fatty acid composition of epididymal plasma has also been determined and compared with blood plasma. By examining the effects of castration, testosterone replacement and cryptorchidism, the accumulation of carnitine and GPC in the epididymis has been shown to be under androgen control. By contrast, the anti-androgens, cyproterone acetate and medroxyprogesterone acetate, did not influence secretory activity, perhaps because their effect was nullified by a high androgen concentration in epididymal plasma. The antifertility compound \ g=a\ \ x=r eq-\ chlorohydrin was also shown to be without effect on the concentration of carnitine and GPC in the reproductive tract. The relationship of carnitine and GPC to the metabolism of fatty acids by spermatozoa is discussed.
Bacillus subtilis 168ts-200B is a temperature-sensitive mutant of B. subtilis 168 which grows as rods at 30 C but as irregular spheres at 45 C. Growth at the nonpermissive temperature resulted in a deficiency of teichoic acid in the cell wall. A decrease in teichoic acid synthesis coupled with the rapid turnover of this polymer led to a progressive loss until less than 20% of the level found in wild-type rods remained in spheres. Extracts of cells grown at 45 C contained amounts of the enzymes involved in the biosynthesis and glucosylation of teichoic acids that were equal to or greater than those found in normal rods. Cell walls of the spheres were deficient also in the endogenous autolytic enzyme (Nacyl muramyl-L-alanine amidase). Genetic analysis of the mutant by PBS1mediated transduction and deoxyribonucleic acid-mediated transformation demonstrated that the lesion responsible for these effects (tag-i) is tightly linked to the genes which regulate the glucosylation of teichoic acid in the midportion of the chromosome of B. subtilis.
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