Saburo Hosomi

Inhibitory effect of 5-phosphoribosyl 1-pyrophosphate and ADP on the nonoxidative pentose phosphate pathway activity

The rate of the conversion of ribose 5-phosphate to hexose 6-phosphates by reaction of the non-oxidative pentose phosphate pathway was measured in the presence of various biological materials. Of 22 compounds tested, PRPP and ADP markedly inhibited the formation of hexose 6-phosphates from ribose 5-phosphate. The transketolase activity in beef liver enzyme preparation was extremely inhibited by PRPP and ADP, but the transaldolase activity was not inhibited. The mode of inhibition of transketolase by PRPP and ADP was a competitive one. The Ki value for PRPP was 0.14 mM and that for ADP 0.54 mM with respect to transketolase. We discuss the possible regulatory roles of ADP and PRPP on pentose phosphate metabolism in the pentose phosphate pathway.

Study on Dihydrodiol Dehydrogenase (II) Modulation of Dihydrodiol Dehydrogenase Activity by Biological Disulfides

Rat liver cytosolic 3α-hydroxysteroid dehydrogenase (EC 1.1.1.50) (dihydrodiol dehydrogenase) which can catalyze the conversion between androsterone and androstanedione in the presence of NADP(H) has also been shown to catalyze the oxidation of benzenedihydrodiol to catechol (Penning et al, 1984, 1985; Hara et al., 1988). From the facts that can oxidize the trans-dihydrodiol of polycyclic aromatic hydrocarbons such as benzo(a)pyrene and benzo(a)anthracene, it has been suggested that dihydrodiol dehydrogenase plays an important role in the detoxification of the polycyclic aromatic hydrocarbon through the effective suppression in the formation of the ultimate carcinogenic anti-diol epoxides (Penning et al., 1984, 1985).

Methods for enzymatic and colorimetric determinations of d-erythrulose (d-tetrulose)

The specific determinations of D-erythrulose by enzymatic assay or colorimetric method, which permit the quantitative determination of between 20 and 400 nmol of the sugar, are described. Enzymatic determination of D-erythrulose made use of the D-erythrulose reductase purified from beef or chicken liver, which catalyzes specifically the reduction of D-erythrulose with concomitant conversion of NADH to NAD+. The colorimetric microdetermination of erythrulose could be carried out by utilizing the phenol-sulfuric acid reaction under low temperature. These methods are simple, rapid, and sensitive, and give reproducible results.

[39] d-erythrulose reductase from beef liver

Publisher Summary This chapter describes the assay method of D-erythrulose reductase isolated from the beef liver. D-erythrulose reductase activity is measured by following the decrease in the absorption of nicotinamide adenine dinucleotide phosphate dehydrogenase [NAD(P)H] at 340 nm, accompanying the reduction of D-erythrulose. The reaction is initiated at 28° by the addition of enzyme. A blank without D-erythrulose must be run to correct for any nonspecific oxidation of NAD(P)H. The steps involved in the purification procedure are (1) homogenization, (2) acetone fractionation, (3) diethylaminoethyl (DEAE)-cellulose chromatography I and II, (4) hydroxyapatite chromatography I and II, and (5) crystallization. D-erythrulose reductase contains 2-3 mol of bound NADP+ per mole of enzyme, so that the extinction coefficient of the enzyme at 280 nm is not constant. Thus, the concentration of pure enzyme protein is calculated from absorbance measurement at 290 nm; at this wavelength, the contribution of NADP+ to the absorbance is negligible. The molecular weight of the enzyme is estimated by sedimentation equilibrium analysis, Sephadex Go200 gel filtration, and sucrose density gradient centrifugation. The subunit molecular weight is determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.