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Biomedical subjects

H R Levy

Publications and source records attributed to H R Levy.

At least 37 records · Page 2Linked to original sources
PubMed1985

Conformations of bound nucleoside triphosphate effectors in aspartate transcarbamylase. Evidence for the London-Schmidt model by transferred nuclear Overhauser effects.

Transferred nuclear Overhauser effects were used to determine the conformations of ATP, CTP, and ITP bound to the regulatory site of aspartate transcarbamylase. The results are in accord with the predictions of the London-Schmidt model [London, R. E., & Schmidt, P. G. (1972) Biochemistry 11, 3136] and show that ATP and CTP bind in the anti conformation while ITP binds in the syn conformation.

Allosteric Site↗
PubMed1984

Glucose-6-phosphate dehydrogenase from rabbit erythrocytes.

Glucose-6-phosphate dehydrogenase (D-glucose-6-phosphate: NADP+ 1-oxidoreductase, EC 1.1.1.49) was purified from rabbit erythrocytes. Initial velocity studies and product and and dead-end inhibitor studies with this enzyme are consistent with a rapid equilibrium random mechanism with an enzyme-NADPH-glucose 6-phosphate dead-end complex.

Animals↗
PubMed1983

A critical appraisal of the effect of oxidized glutathione on hepatic glucose 6-phosphate dehydrogenase activity.

Experiments were undertaken to elucidate the mechanism of the reversal of NADPH inhibition of rat liver glucose 6-phosphate dehydrogenase by oxidized gluthathione alone and in combination with a putative cofactor described by Eggleston & Krebs [(1974) Biochem. J. 138, 425-435]. Evidence is presented that this reversal is largely an artifact, caused by the incorrect application of a control assay procedure and a spurious effect of Zn2+ (added in order to inhibit glutathione reductase) in crude enzyme solutions. When the proper assay procedure is used and glutathione reductase is inhibited with low concentrations of Hg2+, glutathione addition has no effect on NADPH inhibition of glucose 6-phosphate dehydrogenase. No evidence was found for the existence of a cofactor that mediates an effect of glutathione on glucose 6-phosphate dehydrogenase.

Animals↗
PubMed1983

Conformations of nicotinamide coenzymes bound to dehydrogenases determined by transferred nuclear Overhauser effects.

Transferred nuclear Overhauser enhancement was used to examine the conformation of NAD+ and NADP+ bound to glucose-6-phosphate dehydrogenase and glutamate dehydrogenase and of NAD+ bound to lactate dehydrogenase. The results demonstrate that the conformation of the nicotinamide-ribose bond is anti for dehydrogenases with A stereospecificity and syn for dehydrogenases with B stereospecificity. In those dehydrogenases that bind both NAD+ and NADP+, significant differences occur in the conformations of the bound nicotinamide coenzymes.

Animals↗
PubMed1983

Glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides: revised kinetic mechanism and kinetics of ATP inhibition.

The kinetic mechanisms of the NAD- and NADP-linked reactions catalyzed by glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides were examined using product inhibition, dead-end inhibition and alternate substrate experiments. The results are consistent with a steady-state random mechanism for the NAD-linked and an ordered, sequential mechanism with NADP+ binding first for the NADP-linked reaction. Thus, the enzyme can bind NADP+, NAD+, and glucose 6-phosphate, but the enzyme-glucose 6-phosphate complex can react only with NAD+, not with NADP+. This affects the rate equation for the NADP-linked reaction by introducing a term for a dead-end enzyme-glucose 6-phosphate complex. The kinetic mechanisms represent revisions of those proposed previously (C. Olive, M.E. Geroch, and H.R. Levy, 1971, J. Biol. Chem. 246, 2047-2057) and provide a kinetic basis for the regulation of coenzyme utilization of the enzyme by glucose 6-phosphate concentration (H.R. Levy, and G.H. Daouk, 1979, J. Biol. Chem. 254, 4843-4847) and NADPH/NADP+ concentration ratios (H.R. Levy, G.H. Daouk, and M.A. Katopes, 1979, Arch, Biochem. Biophys. 198, 406-413). The kinetic mechanisms were found to be the same at pH 6.2 and pH 7.8. The kinetics of ATP inhibition of the NAD- and NADP-linked reactions were examined at pH 6.2 and pH 7.8. The results are interpreted in terms of ATP addition to binary enzyme-coenzyme and enzyme-glucose 6-phosphate complexes.

Adenosine Triphosphate↗
PubMed1982

Glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides. Isolation and sequence of a peptide containing an essential lysine.

Interaction of glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides with pyridoxal 5'-phosphate and sodium borohydride leads to inactivation and modification of two lysine residues per enzyme dimer that are thought to bind glucose 6-phosphate [Milhausen, M., & Levy, H.R. (1975) Eur. J. Biochem. 50, 453-461]. The amino acid sequence surrounding this lysine residue is reported. Following tryptic hydrolysis of the modified enzyme, two peptides, each containing one pyridoxyllysine residue, were purified to homogeneity and subjected to automated Edman degradation. The sequences revealed that one of these, a heptapeptide, was derived from the other, containing 11 amino acids. Supporting evidence for the role of the modified lysine is provided in the following paper [Haghighi, B., & Levy, H.R. (1982) Biochemistry (second paper of three in this issue)]. End-group analysis of the native enzyme revealed that valine is the N-terminal and glycine the C-terminal amino acid and provides support for the identity of the enzyme's two subunits.

Amino Acid Sequence↗
PubMed1982

Glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides. Kinetics of reassociation and reactivation from inactive subunits.

Glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides is denatured in 8 M urea and dissociated into its two inactive subunits. Denaturation leads to an approximately 80% decrease in protein fluorescence and a 20-nm red shift in the emission maximum. Upon dilution, the urea-treated enzyme regains catalytic activity (approximately 70%). The reactivated enzyme is indistinguishable from the native enzyme based on a number of physicochemical and enzymological criteria. The kinetics of renaturation and reactivation were monitored by measuring the rates of regain of native fluorescence and appearance of activity and the accessibility of histidine residues toward diethyl pyrocarbonate modification. Regain of native fluorescence was too rapid to measure at 25 degrees C; at 5 degrees C the initial phase was also too rapid, but a slower phase was monitored and shown to obey first-order kinetics with k = (5.9 +/- 1.3) x 10(-3) s-1. Reappearance of activity was measured at several protein concentrations; reactivation followed second-order kinetics with k = (4.85 +/- 0.47) x 10(-3) M-1 min-1. Reactivation was stimulated to different degrees by either the initial or delayed addition of NAD+, NADP+, or glucose 6-phosphate. During the initial, rapid phase of renaturation, approximately 3 of the enzyme's 12 histidine residues become unreactive toward diethyl pyrocarbonate; concomitant with the subsequent reactivation, approximately 7 more histidines become inaccessible to diethyl pyrocarbonate. The data are consistent with a model for enzyme renaturation and reactivation in which the unfolded subunits rapidly refold to an inactive structure that can dimerize slowly to generate native enzyme. Specific ligands stimulate reactivation by binding to refolded subunits or incompletely folded dimers.

Glucose-6-Phosphate↗
PubMed1979

Simultaneous analysis of NAD- and NADP-linked activities of dual nucleotide-specific dehydrogenases. Application to Leuconostoc mesenteroides glucose-6-phosphate dehydrogenase.

A method is described which enables one to assay simultaneously the NAD- and NADP-linked reactions of dehydrogenases which can utilize both coenzymes. The method is based on the fact that the thionicotinamide analogs of NADH and NADPH absorb light maximally at 400 nm, a wavelength sufficiently far removed from the absorbance maximum of NADH and NADPH to permit measurements of the simultaneous reduction of NAD+ (or NADP+) and the thionicotinamide analog of NADP+ (or NAD+). Application of the method to glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides reveals differential effects of glucose 6-phosphate concentration on the NAD- and NADP-linked reactions catalyzed by this enzyme which can not be detected by conventional assay procedures and which may have regulatory significance.

Glucosephosphate Dehydrogenase↗
PubMed1979

Glucose-6-phosphate dehydrogenase from lactating rat mammary gland and R3230AC adenocarcinoma.

A number of properties of glucose-6-phosphate dehydrogenase from lactating rat mammary gland and R3230AC rat mammary adenocarcinoma are compared. The main electrophoretic forms of the enzyme from these sources are indistinguishable with respect to charge and molecular weight whereas the minor forms show differences in these properties. The subunit molecular weight and steroid inhibition of the enzymes from the lactating gland and tumor are not significantly different. These results are contrasted with similar studies in mice.

Adenocarcinoma↗