Inhibition of DNA polymerization and DNA transcription to RNA by seminal plasma peptides.

An oligopeptide fraction purified from the extracellular compartment of bull semen and strongly interacting with DNA was shown to hinder mononucleotide polymerizations to DNA and RNA in vitro. The fraction, collectively called seminal plasma inhibitor, was active in the endogenous DNA and RNA polymerase reactions of the nuclei from rat hepatocytes and in the analogous nucleotide polymerizations catalyzed by purified enzymes of bacterial origin. The type of the induced inhibition was studied using the RNA polymerase from Escherichia coli as a representative nucleotidyl transferase. In the enzymatic polycondensation of mononucleotides, the seminal plasma inhibitor appeared to exert its effect mainly by a competitive inhibition for the utilization of DNA templates without specificity with respect to the source and the base sequence of DNA. Concavities of the plots of V0/Vi versus the amounts of inhibitor in the nucleotide polymerizing reactions and of the Dixon plots in the assays of RNA polymerase from E. coli suggested that the isolated oligopeptide fraction contained more than one active molecular species with differential effects at low and high doses. Preliminary results on the microheterogeneity of the seminal plasma inhibitor supported this contention.

Bovine seminal plasma contains a low-molecular-weight factor that inhibits RNA synthesis.

A low-MW factor (800-1000 daltons) extracted from bovine seminal plasma (bSP) and partially purified by a five-step fractionation is very active in inhibiting RNA synthesis by E. coli RNA polymerase with calf thymus DNA as template (70% inhibition at factor:DNA ratio of about 1:100). The same factor also inhibits RNA synthesis in isolated liver nuclei but to a lesser extent. The bSP factor probably exerts its inhibitory activity on initiation rather than on the elongation processes. DNA heat denaturation experiments indicate that the factor stabilizes double-stranded DNA. The activity of bSP factor is almost destroyed by protease (pronase) digestion. Trypsin digestion is ineffective. Consequently, peptide integrity seems to be important for the biological activity. The factor is heat stable and does not contain nucleic acid components. Preliminary analysis indicates the presence of acidic amino acids with no basic or aromatic ones and that the active factor is not a product of histone or protamine degradation. When injected i.p. into 25-day-old female rats, the bSP factor has an inhibinlike activity.

A non-steroidal gametic factor linked to DNA modulates delta-aminolevulinic acid synthase inducibility acting on liver transcriptional and translational processes.

The effect of an acidic factor of low molecular weight (about 1,000 daltons), extracted from bovine spermatozoan DNA, on the inducibility of delta-aminolevulinic acid synthase by ethanol during aging in rat has been examined. The increased enzyme inducibility in 600-day old rats is supported by stimulation of transcriptional and translational processes; on the contrary, in 30-day old rats, the higher enzyme values induced by ethanol are significantly decreased after factor treatment. The active factor is strongly DNA-bound in the native spermatozoan DNA. This would imply a possible role in regulating gene expression in vivo.

Effect of a nonsteroidal gametic factor on senile cataract in the dog.

We have studied the clarifying activity of a purified peptide fraction of gametic origin in senile cataract of the dog. The active substance, administered intramuscularly, had significant and durable clarifying effects on the cortical area of the lens.

Activity of a purified nonsteroidal gametic factor on the inducibility of hepatic delta-aminolevulinic acid synthetase, NADPH-oxidase and tyrosine aminotransferase in senescent rats.

The effect of a water soluble nonsteroidal factor extracted from the male gamete on the activity of certain liver inducible enzymes during aging has been examined. Three enzymes have been studied: delta-aminolevulinic acid synthetase, NADPH-oxidase and tyrosine aminotransferase whose inducibility by ethanol, phenobarbital and ACTH, respectively, show age dependent alterations. The results here reported show that this factor is able to restore the enzyme inducibility in the liver of aging (600-day-old) rats without affecting the response of young (40-day-old) rats. Since the enzyme inducibility is altered during aging, and in the majority of rat hepatomas this factor might enter, possibly, in the regulation of enzyme activity also of neoplastic cells.

Inhibition of gastric acid secretion by a glycoprotein isolated from human urine (human urinary gastric inhibitor).

The gastric antisecretory activity of an inhibitor newly isolated from human urine (Human Urinary Gastric Inhibitor or HUGI) has been studied. HUGI was given intravenously and its activity determined in the following test systems: gastric secretion in the rat with pyloric ligation; gastric secretion in the dog with a Heidenhain pouch stimulated with pentagastrin, histamine and a protein meal; acid secretion by the isolated gastric mucosa of the rat; gastrointestinal motility; bile flow and gall-bladder tone and arterial and venous blood pressure and heart rate. HUGI was found to have marked activity only in the pyloric-ligated rats and in the dogs with Heidenhain pouches stimulated by a protein meal. Particularly in the dog, HUGI (0.1 to 6.4 micrograms/kg, i.v) markedly inhibited gastric secretion, dose-dependently and without changing the plasma gastrin concentration. Negative results were obtained in the other tests, but these results serve to demonstrate that HUGI is an inhibitor well-differentiated from other glycoproteins or peptides with gastric antisecretory activity, such as urogastrone and GIP. The results obtained to date are not sufficient to allow the mechanism of action of HUGI to be defined.

The human urinary gastric inhibitor displays blood-group activity.

The human urinary glycoprotein with gastric antisecretory activity displays a marked blood-group activity corresponding to the blood-group of the subject examined. Several studies exclude the possibility that the activity is due to contaminants.

Effect of the inhibin-like factor from bull spermatozoa on sexual maturation of the male rat.

The s.c. administration for 25 days of the inhibin-like factor extracted from bull spermatozoa to immature 20-day-old male rats reduced serum FSH levels by about 60% and serum LH and testosterone levels by about 50%. However, neither tubular nor interstitial modifications of the histological appearance of the testis were found and there were no changes in the activity of delta 5-3 beta-hydroxysteroid dehydrogenase. The assay described could be usefully employed for evaluating the activities of inhibin-like factors.

Chemical and physical properties of the human urinary glycoprotein with gastric antisecretory activity.

The chemical and physical properties of the high-molecular-weight glycoprotein (SO20, w = 8S; Ve=Vo on Sephadex G-200) with gastric antisecretory activity extracted from the urine of pregnant women were studied. Gel filtration in the presence of sodium dodecyl sulphate and sodium dodecyl sulphate/polyacrylamide-disc-gel electrophoresis indicated subunit mol.wts. of 16 000 +/- 1500 and 13 000 +/- 1000 respectively. Reaggregation of the subunits and partial recovery of the biological activity were observed on removal of the detergent. The partial C-terminal sequence was found to be Phe-Tyr-Leu-Val-OH, whereas glycine appears to be the N-terminal amino acid. The carbohydrate composition was examined; all galactosamine was found to be O-glycosidically linked to the polypeptide chain.