Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.26.9 (ribonuclease)
6,589 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The presence of a nuclear DNA polymerase in mouse sperm from adult testes has been confirmed and the properties of this enzyme further investigated. This activity was shown to be greatly enhanced by treating the spermatozoa with methanol or ethanol before incubation in the reaction medium or by their addition in small amounts to this medium. It was protected against degradation by nuclear proteases by adding soybean trypsin inhibitor and was stimulated by ATP. It was found to be Mg2+ dependent (optimum concentration: 7.5 mM), DNA dependent, and all four deoxynucleoside triphosphates were needed for optimal reaction. The radioactive acid-precipitable product of polymerization was not eliminated by organic solvents, nor by pronase, ribonuclease or by nuclease S1; however, it was converted to a large extent to acid-soluble products by pancreatic deoxyribonuclease. Since it was only partially solubilized by Triton X-100, it therefore did not appear to be preferentially associated with the nuclear membranes. The activity recovered after incubation depended also on the pH (optimum at pH 8.3) and did not work well in a medium for DNA polymerase alpha. The temperature for maximum incorporation of nucleotides was found to be 32 degrees C and, under our conditions, the reaction was linear for 30 min. The DNA polymerase activity was inhibited by low and high concentrations of KCl. It was not lowered by N-ethylmaleimide or p-hydroxymercuribenzoate; urea slightly stimulated the reaction and this stimulation was reversed by subsequent treatment with N-ethylmaleimide. Actinomycin D (40 mug/ml), ethidium bromide (25--50 muM), netropsin (5--50 mug/ml), and spermidine (0.5--2.5 mM) lowered the polymerization of DNA precursors. The nuclear enzyme could shift from the endogenous template to activated exogenous calf thymus DNA, the resulting nuclear radioactivity being reduced. The endogenous DNP template ability was not increased by deoxyribonuclease activation according to the method of Aposhian and Kornberg (J. Biol. Chem. (1962) 237, 519--525) suggesting that the amount of DNA polymerase associated with chromatin was probably limiting the reaction. The DNA polymerase activity detected in mouse sperm nuclei has numerous properties of low molecular weight DNA polymerases (DNA polymerase beta) reported in several eukaryotic organisms.
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PMID:Further characterization of a DNA polymerase activity in mouse sperm nuclei. 1 3

Under defined conditions, in the presence of 10 mg/ml of bovine serum albumin, cauda epididymal rat spermatozoa displayed vigorous motility, and a high proportion (81%) of eggs were fertilized. In contrast, no fertilization was observed after omission of albumin, or replacement of the protein by 10 mg/ml of cytochrome c, beta-globulin, gamma-globulin, hemoglobin, lysozyme, and polyvinylpyrrolidone, and 5 mg/ml of ribonuclease. However, high motility occurred in suspensions containing 3 x 10(6) spermatozoa/0.1 ml of medium with cytochrome c, beta-globulin, or gamma-globulin. In medium with 1 mg/ml of ovalbumin, 7% (2/29) eggs were fertilized. Use of defatted albumin resulted in a higher rate of fertilization than unmodified albumin (87 vs 70%), and this difference approached statistical significance. No fertilization was obtained in the presence of albumin presaturated with cholesterol. These results suggest that: (a) rat sperm cells failed to capacitate in the absence of albumin; (b) the protein exerted more than a nonspecific macromolecular effect; and (c) lipids associated with albumin may modify its ability to promote sperm capacitation.
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PMID:Influence of serum albumin on the fertilizing ability in vitro of rat spermatozoa. 125 Aug 65

The pattern of proteins in the soluble fraction of the cytoplasm of the rat epididymis was studied by acrylamide gel electrophoresis. The components of five distinct bands, labelled A, B, C, D and E, were found to be sensitive to changes in androgen in the blood. Castration for 14 days produced a sharp decrease in the colour intensity of bands B-E when stained with Amido black. After 21 days of castration, bands D and E were undetectable, bands B and C were severely diminished and band A was more intense. Seven days of replacement with testosterone (1 mg/day) induced a return towards a normal pattern. The degree of restoration was inversely proportional to the duration of castration. Quantitation by densitometry showed that the relative contributions of bands B-E to the region A-E were 61% in the control rat, only 27% after 21 days of castration and 35% when testosterone was given between days 14 and 21 of castration. The components of bands A-E are presumed to be proteins since the electrophoretic pattern was altered by digestion with pronase but not by ribonuclease, phospholipase C or neuraminidase. Epididymides from castrated and androgen-treated castrated rats were incubated with 14C- and 3H-labelled mixed amino acids respectively. After co-electrophoresis the ratio 3H: 14C rose from a baseline of 2-5 in band B, 32 in band C and 7 in bands D and E. Molecular weights were estimated as 27900 for B, 23100 for C and 34400 for D. Band A had the same electrophoretic mobility as serum albumin. Bands B and C were also present in testicular cytosol. Bands D and E were only found in the epididymis, localized mainly within the lumen of the tubules. Bands B-E increased with age during sexual maturation, bands D and E became detectable in the 20-day-old rats. Preliminary evidence indicates that the proteins in bands C, D and E can be removed from caput spermatozoa by washing.
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PMID:Androgen-controlled specific proteins in rat epididymis. 127 Sep 59

Spermatozoa from the cauda of the epididymis of the hamster and rat were incubated with [5-(3)H]uridine and glucose. By using a procedure avoiding bacterial and other cellular contamination, sonic extracts were prepared and digested with deoxyribonuclease and Pronase. Radioactive RNA of high molecular weight was isolated by two methods: (a) gel filtration on Sephadex G-75 columns and (b) polyacrylamide-gel electrophoresis in which it migrated in the region of 28S and 23S RNA markers. The macromolecules were alkali-labile and hydrolysed by ribonuclease. From (3)H radioactivity and E(260) of the isolated RNA the rate of incorporation of uridine into RNA of spermatozoa was calculated to be 0.1-0.5nmol/h per mg of RNA.
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PMID:Ribonucleic acid synthesis by spermatozoa from the rat and hamster. 474 26

Human seminal fluid, at low dilutions, prevented the binding of aggregated human IgG (AHG) to bull spermatozoa. Seminal fluids from vasectomized men were also inhibitory. Preincubation of the seminal fluid with the spermatozoa prior to washing and addition to AHG had no inhibitory effect, indicating that the fluid component was reacting directly with AHG. Human seminal fluid was fractionated by gel exclusion chromatography on Ultrogel AcA-34, and AHG inhibitory activity was found in fractions corresponding to a molecular weight of 94,000. The activity in this fraction was stable to boiling for 10 min. It was sensitive to pronase but resistant to glycosidase, phospholipase C, neuraminidase, ribonuclease, and deoxyribonuclease, indicating that it was a protein. The gel filtration fraction readily bound recrystallized Fc and AHG; IgG was bound to a lesser extent, and no reactivity was observed with F(ab')2, IgA, or IgM. Thus, the seminal fluid fraction appeared to specifically react with the Fc portion of IgG. The seminal fluid Fc-binding protein was isolated by affinity chromatography on Fc coupled to CNBr-activated Sepharose 4B. Scatchard analysis revealed that the binding of the seminal fluid Fc-binding protein to recrystallized Fc is reversible and had a Kd of approximately 3 x 10(-6) M.
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PMID:An IgG-Fc binding protein in seminal fluid. 622 60

The native dimer bovine seminal ribonuclease, AS RNase, and its four modified derivatives (carboxymethylated, succinylated, oxidated and reduced) were examined for their effects on PHA- or PWM-stimulated human lymphocytes, mixed lymphocyte cultures and lymphoblastoid cell line line (Molt-3, RAJI, UHKT-2 and UHKT-5). Among all substances tested the native AS RNase exerted the strongest suppressive effect on PHA-, PWM-, and MLR- stimulated lymphocytes. At concentrations of 5 and 100 microgram/ml AS RNase inhibited lymphocyte stimulation in MLR by 40 and 95%. The carboxymethylated and reduced derivatives possessed inhibitory effect one order lower, while the succinylated derivative showed a negligible effect, and the oxidated derivative was ineffective. The inhibitory effect on PHA-stimulated lymphocytes was not reduced when AS RNase was added to lymphocyte culture 24 or 48 h later. Likewise, the 20-fold increase in PHA concentration did not cause any decrease in inhibitory effect of AS RNase. AS RNase added to transformed lymphocytes simultaneously with 3H-TdR did not affect the thymidine uptake. As RNase also had the strongest inhibitory effect on growth, viability and the DNA synthesis in all lymphoblastoid cell lines tested. After 72 h cultivation with AS RNase at a concentration of 100 microgram/ml, the DNA synthesis decreased by more than 90% in Molt-3 and UHKT-2 cells. Simultaneously reduction in cell count and increased numbers of dead cells compared to control cultures were found. Carboxymethylated derivative at the same concentration inhibited thymidine incorporation into the DNA by 70% but did not cause any cytotoxic effect. Other derivatives showed negligible or null effects. These results are in agreement with some data published earlier, that the dimer of seminal ribonuclease exerts a significant antitumour effect. The immunosuppressive effects of AS RNase observed in the present paper suggest that this compound is one of the components of seminal plasma which may take part in suppression of immune responses of the female reproductive tract to spermatozoa.
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PMID:Inhibitory effect of bovine seminal ribonuclease on activated lymphocyte and lymphoblastoid cell lines in vitro. 645 59

Wistar male rats received an intratesticular injection (at 114 and 265 days of age) of 3 mg of partially purified bull seminal ribonuclease (AS RNase) or saline. It was found that sexual behaviour (initiation of copulation as well as copulatory behavioural pattern) of experimental males was not changed, but the ability of these males to fertilize females was evidently suppressed. In addition to significantly lower weights of testes and epididymis, inhibition of seminiferous epithelium development (aspermatogenesis) associated with the absence of spermatozoa was determined in cauda epididymidis in experimental animals. However, Leydig cells remained without changes. Plasma testosterone levels of AS RNase treated males were not altered in comparison with the controls. Thus AS RNase specifically impaired spermatogenesis but did not influence androgen action and sexual behaviour.
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PMID:The effect of bull seminal ribonuclease on reproductive organs and sexual behaviour in male rats. 645 96

Exogenous calf thymus whole histones showed a high degree of specificity to cause agglutination of rat epididymal spermatozoa. Histones had markedly greater (approximately 5-fold) agglutination activity than did salmon protamine whereas a variety of proteins, including strongly basic ones such as herring protamine sulphate, ribonuclease, cytochrome C and lysozyme, had no detectable agglutination activity. Histones F-3 and F-2a had the greatest activity for cell agglutination. Polyamines (5 mM), sialic acid (5 mM) and basic or acidic amino acids (10 mM) had no effect on histone (approximately 8 microM)-mediated sperm agglutination. 32P-Labelled histones showed a high specificity for binding to intact spermatozoa. The binding was saturable at a histone concentration of approximately 0.3 mg/ml and nearly completely displaced at saturating concentrations of native histones. Only unlabelled protamines competed to a small extent for binding of 32P-labelled histones to spermatozoa. The data are consistent with the view that histones bind specifically to sperm surface receptor sites before agglutination of cells.
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PMID:Histone-mediated agglutination of epididymal spermatozoa and the occurrence of histone receptors on the rat sperm surface. 725 26

Bovine seminal plasma (BSP) contains four similar acidic proteins, previously designated as BSP-A1, BSP-A2, BSP-A3, and BSP-30-kDa. These proteins are secreted by the seminal vesicles and coat the surface of the spermatozoa after ejaculation. The binding site of BSP proteins on the sperm surface has been identified as choline phospholipids on the plasma membrane. This study was undertaken to determine whether BSP proteins modulate capacitation of bovine spermatozoa induced by heparin. Bovine epididymal spermatozoa were washed and incubated in buffer containing BSP proteins and then washed and incubated with heparin. The percentage of capacitated spermatozoa was determined under the microscope after the acrosome reaction has been initiated with the addition of lysophosphatidylcholine. The results demonstrated that epididymal sperm undergo the acrosome reaction only in the presence of BSP proteins. This effect was concentration-dependent and reached a maximum level of a 3-5-fold increase at 20-40 micrograms/ml BSP protein concentrations. In contrast, ribonuclease (purified from bovine seminal fluid) or seminal fluid proteins depleted of BSP proteins (by sequential absorption of BSP proteins on gelatin-Agarose and DEAE-Sephadex columns) showed no significant potentiating activity. The purified BSP proteins were more active than crude alcohol precipitates of bovine seminal plasma. These results indicate that BSP proteins are regulatory factors of capacitation.
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PMID:Phosphatidylcholine-binding proteins of bovine seminal plasma modulate capacitation of spermatozoa by heparin. 763 45

Striped bass are seasonal breeding fish, spawning once a year during the spring. All 3-yr-old males are sexually mature; however, 60-64% of the fish mature earlier as 1- or 2-yr-old animals. The endocrine basis underlying early maturity in 2-yr-old males was studied at the molecular level by monitoring changes in pituitary beta FSH and beta LH mRNA levels by ribonuclease protection assay, and correlating these changes to stages of testicular development. In maturing males, the mRNA levels of beta FSH were elevated during early spermatogenesis, whereas beta LH mRNA levels peaked during spermiation. The appearance of spermatozoa in the testis was associated with a decrease in beta FSH mRNA and a rise in beta LH mRNA abundance. Immature males had lower levels of beta LH mRNA than maturing males, but there were no differences in beta FSH mRNA levels between immature and maturing males. The regulation of gonadotropin gene expression in 2-yr-old males was studied by the chronic administration of GnRH analogue (GnRHa) and testosterone (T), with or without pimozide (P) supplementation. In immature males, the combination of T and GnRHa stimulated a three- to fivefold increase in beta FSH and beta LH mRNA levels, but the same treatment had no effect on gonadotropin gene expression in maturing males. In addition, the coadministration of P to immature males suppressed the stimulatory effect of GnRHa and T on beta FSH and beta LH mRNA levels, suggesting that dopamine may have a novel role in regulating gonadotropin gene expression.
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PMID:Early maturity in the male striped bass, Morone saxatilis: follicle-stimulating hormone and luteinizing hormone gene expression and their regulation by gonadotropin-releasing hormone analogue and testosterone. 1109 Apr 37


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