T.A. Venkitasubramanian

Effect of aflatoxins on oxidative phosphorylation by rat liver mitochondria.

The in vitro effect of aflatoxins M1, B1 and G1 on oxidative phosphorylation by rat liver mitochondria with succinate as substrate has been studied. All these toxins inhibit the electron transport chain at a 1-10-4 M concentration and the site of inhibition is between cytochrome b and cytochrome c or c1. Aflatoxin M1 (AFM1) uncouples oxidative phosphorylation at a concentration of 1-10-6 M and reduces the ADP:O ratio, whereas aflatoxin B1 (AFB1) at 1-10-6 M concentration uncouples oxidative phosphorulation but does not affect the ADP:O ratio. At a concentration of 1-10-5 M, AFB1 also decreases the ADP:O ratio along with the uncoupling of oxidative phosphorylation. Aflatoxin G1 (AFG1) acts as an uncoupler at a relatively higher concentration of 1-10-4 M. Preincubation of mitochondria with these aflatoxins resulted in inhibition of respiration and uncoupling of rat liver mitochondria.

Metabolism of mycobacteria

The metabolism of mycobacteria have been studied with reference to carbohydrate, lipids, nitrogen metabolism and oxidative phosphorylation. Some of the enzymes of glycolytic pathway, tricarboxylic acid cycle and lypogenic enzymes were purified, characterized and their kinetic properties investigated. The effect of age of the culture and environmental factors on different aspects of metabolism of mycobacteria were also studied. A comparison of lipid profile in various species of mycobacteria grown in different culture conditions were made. The metabolism of spheroplasts isolated from mycobacteria has been established with respect to their energy charge and to synthesize peptidoglycan using D-alanine as the precursor

Fructose diphosphate aldolase from Mycobacterium smegmatis: Purification and properties☆☆☆

Abstract Fructose diphosphate aldolase has been purified to homogeneity from Mycobacterium smegmatis. Physicochemical studies showed that the enzyme is a tetramer of molecular weight 158,000. Mycobacterium smegmatis aldolase, though a bacterial enzyme, possesses properties similar to other class I aldolases. Inactivation of the enzyme by sodium borohydride in presence of dihydroxyacetone phosphate suggested the formation of a Schiff-base intermediate.

Early effects of excessive retinol intake on gluconeogenesis: Involvement of adrenals in the increased activities of gluconeogenic enzymes of rat liver☆

A stimulation of gluconeogenesis by excessive intake of retinol is suggested on the basis of enhanced incorporation of [2-14C]glycine into liver glycogen in rats fed excess retinol. Among the key gluconeogenic enzymes studied, activities of hepatic PEP-carboxykinase, fructose-1,6-bisphosphatase, glucose-6-phosphatase, and alanine aminotransferase were markedly increased, whereas that of pyruvate carboxylase remained unaltered. However, feeding of retinol to bilaterally adrenalectomized rats or rats treated with actinomycin D failed to cause significant increase in the activities of these enzymes. It is concluded that (i) gluconeogenesis is stimulated by excess retinol due to, perhaps, increased activities of key gluconeogenic enzymes, and (ii) adrenals are directly or indirectly involved in the retinol-mediated increase in the activities of the gluconeogenic enzymes. Also, data are presented that indicate a requirement for protein synthesis for the expression of retinol-mediated alterations in the activities of gluconeogenic enzymes.

Fructose diphosphate aldolase from Mycobacterium smegmatis: Functional similarities with rabbit muscle aldolase☆☆☆

Abstract Fructose diphosphate aldolase of Mycobacterium smegmatis is found to be a class I type aldolase and possesses functional similarities with rabbit muscle aldolase with respect to the amino acid residues at the catalytic site. The presence of a lysine residue at the active site is indicated by the formation of a Schiff-base with the substrate. The lower degree of inactivation compared to rabbit muscle aldolase on treatment with carboxypeptidase-A suggests the absence of an essential terminal tyrosine residue. Participation of histidine residues in enzyme catalysis is suggested by the photoinactivation of the enzyme in presence of methylene blue. Finally, thiol groups do not seem to have a direct role in catalysis.

Oxidases in Aspergillus parasiticus in relation to aflatoxin biosynthesis

Abstract The effects of asparagine and zinc were studied on the nucleotide oxidases and lipid peroxidase during secondary biosynthesis in A. parasiticus . The activity of NADPH and NADH oxidases was higher in zinc deficient cultures as compared to those in sucrose-low salt cultures and under asparagine deficiency, thereby lowering the NADPH/NADP and NADH/NAD ratios. Lipid peroxidation was increased under conditions of reduced aflatoxin production probably because of greater availability of NADPH. Lower activities of lipid peroxidases and nucleotide oxidases may be favourable for optimum aflatoxin production.

Induction of mixed function oxidases on oral administration of dieldrin

The administration of dieldrin (30 mg/kg body weight) caused an increase in the liver weight of rats. The metabolism of aflatoxins B1 and G1 by the microsomes obtained from the liver of dieldrin-treated animals was enhanced significantly as compared to the controls showing that dieldrin increased the activity of mixed function hydroxylases. Dieldrin caused an increase in the activity of liver microsomal NADPH oxidase and a decrease in the lipid peroxidation. Dieldrin brought about an increase in the phosphatidylcholine content of rat liver.

Dieldrin toxicity and in vivo incorporation of DL-[1-14C]leucine

The in vivo effect of a single oral dose (30mg/kg body weight) of dieldrin on proteolipid and phosphatidopeptide content of liver and brain and on total protein of liver, brain, plasma, muscle and kidney of rat was studied. Incorporation of (14C)leucine into total protein of liver was increased whereas labelling of total protein of muscle was decreased. Labelling of total protein of other tissues was unchanged. Incorporation into liver phosphatidopeptides was increased and this was consistent with an involvement of group. Proteolipid protein content of brain was increased and that of liver unchanged. There was, however, no change in the labelling of brain or liver proteolipids.

Metabolic Potential of the Adipose Tissue of Rats during Hyper- and Hypovitaminosis A

Abstract Effect of excess feeding and depletion of vitamin A on the ability of adipose tissue to maintain plasma free fatty acid levels has been studied in rats. Both in hypervitaminosis A (fed 9 mg of retinol for 2 consecutive days) and in vitamin A deficiency (kept on a vitamin A-deficient diet for 6 weeks) the rats showed elevated levels of plasma free fatty acids. Hypervitaminosis A caused a decrease in the fatty acid release from adipose tissue on in vitro incubation, probably due to lowered levels of cyclic AMP. On the other hand, adipose tissue from vitamin A-deficient animals showed an increased lipolytic rate as compared to that of the controls. No change in the lipogenic ability was indicated in either of the conditions as indicated by the activities of enzymes involved in this process. We conclude that the fatty acid homeostasis can be greatly influenced by the vitamin A status of the animals.

Purification and properties of pyruvate kinase from Mycobacterium smegmatis

Pyruvate kinase (ATP:pyruvate 2-O-phosphotransferase, EC 2.7.1.40) from Mycobacterium smegmatis has been purified to homogeneity through a seven-step procedure with a yield of 16% and specific activity of 220 units/mg protein. The purified enzyme had a molecular weight of 230,700 and was composed of four subunits with identical molecular weights of 57,540. Analysis of amino acid composition revealed a low content of aromatic amino acids. The enzyme exhibited sigmoidal kinetics of varying concentrations of phosphoenolpyruvate, the degree of cooperativity and S0.5v value for phosphoenolpyruvate being strongly dependent on the pH of the reaction mixture. Among the nucleoside diphosphates acting as substrate for pyruvate kinase, ADP was the best phosphate acceptor, as judged by its lowest Km value. The enzyme showed an absolute requirement for divalent cations (either Mg2+ or Mn2+), but monovalent cations were not necessary for activity. Other divalent cations inhibited the Mg2+-activated enzyme to varying degrees (Ni2+ greater than Zn2+ greater than Cu2+ greater than Ca2+ greater than Ba2+). The differences in the kinetic responses of the enzyme to Mg2+ and Mn2+ are discussed.