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Biomedical subjects

E Kun

Publications and source records attributed to E Kun.

At least 73 records · Page 4Linked to original sources
PubMed1983

Spectral analysis of the conformation of polyadenosine diphosphoribose. Evidence indicating secondary structure.

Based on boronic acid chromatography, a rapid method was developed for the purification of polyadenosine diphosphoribose oligomers of various chain lengths. Polyadenosine diphosphoribose oligomers obtained by the new method were distinguished on the basis of spectral criteria. The purity of polyadenosine diphosphoribose was determined by high performance liquid chromatography, which is a highly sensitive method for the detection of contamination by nucleotides derived from other nucleic acids. The A280/A260 ratio, which has been used in the past as one of the criteria of purity of oligomers, was found to be an unsuitable index of purity because it significantly varied as a function of temperature or ionic strength, exhibiting characteristics of temperature- and ionic strength-dependent hypochromicity. Hypochromicity was highly significant at 260 nm but not at 280 nm. The A280/A260 ratio showed a correlation with the oligomer chain length, significantly diminishing below a chain length of 9 adenosine diphosphoribose units, with concomitant loss of hypochromicity. The increase in A280/A260 ratio above a chain length of nine was small and gradual. Circular dichroism of long, medium, and short oligomers was determined at varying temperatures (5-72 degrees C). For long chains, a temperature-dependent change of the major negative theta value and a red shift occurred from 249.5 to 267.5 nm. With medium chain length oligomers, there was no decrease in the theta value at 249.5 nm, only a red shift. In the case of short chain oligomers the major negative theta value was at 267.5 nm and its position was temperature-independent with only a small temperature-related decrease in size. At 72 degrees C, the different patterns of circular dichroism of the three groups of oligomers of differing chain lengths became indistinguishable. These results were interpreted to indicate a significant secondary structure of polyadenosine diphosphoribose of long chain length.

Chromatography, High Pressure Liquid↗
PubMed1983

Cell cycle-dependent intervention by benzamide of carcinogen-induced neoplastic transformation and in vitro poly(ADP-ribosyl)ation of nuclear proteins in human fibroblasts.

Human fibroblasts were subjected to nutritionally induced G1 block, followed by release and subsequent entry into S phase, and exposed to nontoxic concentrations of carcinogens in early S phase. Cell transformation occurred as determined by early morphologic cell alterations, anchorage-independent colony formation, cell invasiveness, and augmentation of Ab 376 human malignancy-specific cell-surface antigenic determinant. Methylazoxymethanol acetate was the most potent transforming agent at doses that were negative in toxicity tests. Benzamide (10 microM intracellular concentration), a specific inhibitor of poly(ADP-ribose) polymerase, prevented transformation in a cell cycle-specific manner, maximal prevention coinciding with early S phase, also characteristic of maximal susceptibility to transformation. Neither an interference of carcinogen deoxyguanosine nucleoside adduct formation nor a chemical reaction between benzamide and carcinogens was detected. Methylazoxymethanol acetate at transforming but nontoxic dose partially inhibited poly(ADP-ribosyl)ation to about the same extent as benzamide. However, simultaneous exposure of cells to both agents in early S phase, resulting in the prevention of transformation, augmented poly(ADP-ribosyl)ation above the controls. Enzymatic activities ran parallel with the formation of DNA-associating polymer-nonhistone protein adducts that are assumed to regulate the physiological function of chromatin at the structural level.

Antigens, Neoplasm↗
PubMed1982

Steric inhibition of phenylboronate complex formation of 2'-(5"-phosphoribosyl)-5'-AMP.

Unlike all other adenine nucleotides which contain cis-diols in their ribose groups, the degradation product of polyadenosine diphosphoribose, 2'-(5"-phosphoribosyl)-5'-AMP, does not bind to boronate affinity columns. This anomalous behavior occurs even under conditions of high ionic strength (1 M salt), indicating that the absence of binding is not due to generalized repulsion between the negatively charged phosphate groups of 2'-(5"-phosphoribosyl)-5'-AMP and the negatively charged boronate sites of the resin. Despite the lack of boronate binding the presence of a chemically functional cis-diol group in 2'-(5"-phosphoribosyl)-5'-AMP was verified by quantitative periodate oxidation. Subsequent dephosphorylation rendered the cis-diol in ribosyladenosine accessible to binding to the boronate sites; therefore, one or both of the phosphate groups sterically interfere with complexation of the cis-diol with boronate. Computer model-building studies based on NMR data implicate the 5'-phosphate group of the molecule, is located sterically near the cis-diol group, inhibiting the complex formation with boronate.

Adenosine Monophosphate↗
PubMed1981

Quantitative isolation of oligo- and polyadenosine-diphosphoribosylated proteins by affinity chromatography from livers of normal and dimethylnitrosamine-treated Syrian hamsters. In vivo and in vitro metabolism of the homopolymer.

Polyadenosine- and adenosine-diphosphoribosylated proteins of hamster liver were quantitatively isolated with the aid of m-aminophenyl boronic acid glutaryl hydrazide polyacrylamide affinity resin by selective adsorption at pH 8.2 and elution at pH 4.0. Polymer-free proteins, DNA, and RNA are readily separated from adenosine-diphosphoribosylated proteins. The total quantity of proteins that is covalently modified by the homopolymer is 14.3 micrograms/mg of DNA or 37.4 micrograms/g of liver in controls and 38.7 micrograms/mg of DNA or 116 micrograms/g of liver in dimethylnitrosamine-treated hamsters. Polymer content increases from 9 to 15 nmol/mg of DNA to 42 to 118 nmol/mg of DNA following treatment with dimethylnitrosamine. Pulse labeling with [14C]ribose results in a parallel doubling in dimethylnitrosamine-treated animals of the specific activities of adenosine- diphosphoribose and NAD+ and of the [14C]ribose content of polyadenosine-diphosphoribose of chain length between 20 and 40, indicating chain elongation of pre-existing larger polymers. Two groups of proteins that are isolated as polyadenosine-diphosphoribose adducts are increased significantly after treatment with dimethylnitrosamine, one minor component of a mass between 100-112 X 10(3) daltons, and a major group exhibiting a mass of 158-162 X 10(3) daltons. Polyadenosine-diphosphoribose synthetase activity of isolated hepatic nuclei is increased by 32-37% after dimethylnitrosamine treatment, and since the change in glycohydrolase activity is negligible relative to the increase in synthetase, the augmentation of polyadenosine-diphosphoribosylated proteins can be explained by the increased synthetase of nuclei. The molecular size distribution of DNA in liver nuclei of control and dimethylnitrosamine-treated hamsters is indistinguishable.

Adenosine Diphosphate Ribose↗
PubMed1981

Age-dependent variation of rates of polyadenosine-diphosphoribose synthesis by cardiocyte nuclei and the lack of correlation of enzymatic activity with macromolecular size distribution of DNA.

Cardiocyte nuclei from neonate (5-day-old) and from adult (90-day-old) male Wistar rats were selectively isolated by a technique that eliminates interference by noncardiocyte nuclei (Jackowski, G., and Liew, C. C. (1980) Biochem. J. 188, 363-373). Polyadenosine-diphosphoribose synthetase activity of cardiocyte nuclei of neonates was ten times greater than in nuclei of adult rats, as calculated from initial velocity linear rate measurements. The molecular size of DNA extracted from cardiocyte nuclei of neonates was significantly larger (peak at 54 S) than DNA of cardiocyte nuclei of adults (peak at 33 S) as determined by alkaline sucrose density gradient ultracentrifugation (Knopf, K. W., and Weissbach, A. (1977) Biochemistry 16, 3190-3194). Comparison of the molecular sizes of DNA extracted from whole cardiac tissue with DNA isolated from cardiocyte nuclei shows that no DNA fragmentation takes place during the process of isolation of nuclei. Polyadenosine-diphosphoribose glycohydrolase activity was about 30% higher in cardiocyte nuclei of neonates than in adults, but the activity represents only 10% of the rate of the synthetase. Results demonstrate that polyadenosine-diphosphoribose metabolism in differentiated tissues is not directly related to the molecular size of DNA.

Aging↗
PubMed1981

Tubular proteinuria after perinatal hypoxia.

Urinary total protein (UTP) and urinary protein pattern have been studied in 23 newborn infants with Apgar scores less than or equal to 3 at one minute or acidosis (pH less than or equal to 7.15) on the first day. On the first and second day UTP excretion was increased in 13 out of 18 patients. At this time the excretion of low molecular weight microproteins (T-4 and T-5) was elevated in 12 patients without increased plasma urea concentration in any case. The increased excretion of the smallest microproteins T-4/T-5 is an early sign of an impaired tubular function.

Acidosis↗
PubMed1979

The in vivo distribution of immunoreactive larger than tetrameric polyadenosine diphosphoribose in histone and non-histone protein fractions of rat liver.

The macromolecular association of immunoreactive naturally occurring polyadenosine diphosphoribose n greater than 4 with histones and non-histone proteins was determined with the aid of an improved method of extraction of polyadenosine diphosphoribose and a combination of radioimmunoassay and molecular filtration. More than 99% of the naturally occurring polyadenosine diphosphoribose n greater than 4 was present in rat liver in covalent association with non-histone proteins. The chain length of the polymer varied between n = 4 and n = 34. Less than 1% of naturally occurring polyadenosine diphosphoribose n greater than 4 was almost evenly distributed between histone fractions f1, f2a, f2b, and f3. Adenosine diphosphoribose polymers of relatively long chain length were also detected in the histone fractions. The covalent association of polyadenosine diphosphoribose with non-histone proteins was demonstrated by affinity chromatography.

Animals↗