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Carnitine palmitoyltransferase 1 (CPT1) catalyzes the conversion of palmitoyl-CoA to palmitoylcarnitine in the presence of L-carnitine, thus facilitating the entry of fatty acids to mitochondria, in a process that is physiologically inhibited by malonyl-CoA. To examine the mechanism of CPT1 liver isoform (CPT1A) inhibition by malonyl-CoA, we constructed an in silico model of both its NH 2 -and COOH-terminal domains. Two malonyl-CoA binding sites were found. One of these, the "CoA site" or "A site," is involved in the interactions between NH 2 -and COOH-terminal domains and shares the acyl-CoA hemitunnel. The other, the "opposite-to-CoA site" or "O site," is on the opposite side of the enzyme, in the catalytic channel. The two sites share the carnitine-binding locus. To prevent the interaction between NH 2 -and COOHterminal regions, we produced CPT1A E26K and K561E mutants. A double mutant E26K/K561E (swap), which was expected to conserve the interaction, was also produced. Inhibition assays showed a 12-fold decrease in the sensitivity (IC 50 ) toward malonyl-CoA for CPT1A E26K and K561E single mutants, whereas swap mutant reverts to wild-type IC 50 value. We conclude that structural interaction between both domains is critical for enzyme sensitivity to malonyl-CoA inhibition at the "A site." The location of the "O site" for malonyl-CoA binding was supported by inhibition assays of expressed R243T mutant. The model is also sustained by kinetic experiments that indicated linear mixed type malonyl-CoA inhibition for carnitine. Malonyl-CoA alters the affinity of carnitine, and there appears to be an exponential inverse relation between carnitine K m and malonyl-CoA IC 50 . Carnitine palmitoyltransferase 1 (CPT1)4 catalyzes the conversion of long-chain fatty acyl-CoAs to acylcarnitines in the presence of L-carnitine. This is the first step in the transport of long-chain fatty acids from the cytoplasm to the mitochondrial matrix, where they undergo ␤-oxidation. CPT1 is tightly regulated by its physiological inhibitor malonyl-CoA. This regulation allows CPT1 to signal the availability of lipid and carbohydrate fuels to the cell (1). Mammalian tissues express three isoforms: CPT1A (liver), CPT1B (muscle and heart), and CPT1C (brain), which are the products of different genes (2-4). CPT1A and -B have 62% amino acid identity, but they are differentially regulated by malonyl-CoA. CPT1A is inhibited to a much lesser extent than CPT1B, which may explain why fatty acid oxidation is more finely regulated in the heart than in the liver. CPT1 is a potential target for the treatment of metabolic disorders involving diabetes and coronary heart disease (5). The interaction between malonyl-CoA and CPT1C may be involved in the "malonyl-CoA signal" in hypothalamic neurons regulating food intake and peripheral energy expenditure (6).It has been postulated that there are two malonyl-CoA binding sites in the molecule of CPT1A (7,8). Kinetic studies indicate that there is a high affinity binding site and a low affinity binding site (9 -13...

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Repression of the synthesis of isocitrate lyase by glucose and/or induction of the synthesis of isocitrate lyase by acetate in Phycomyces blakesleeanus were demonstrated. Both glycerol and ethanol failed to induce isocitrate lyase activity. Furthermore, glucose appeared to cause an in vivo catabolite inactivation of the derepressed enzyme. Isocitrate lyase was inactivated both reversibly and irreversibly by glucose.

This study investigated the antimicrobial activity of 3 natural (thymol, carvacrol, and gallic acid) and 2 synthetic [butylated hydroxyanisole (BHA) and octyl gallate] phenolic compounds, individually and in binary combinations, on 4 dairy isolates of Enterococcus faecalis with different virulence factors (β-hemolytic, gelatinase, or trypsin activities; acquired resistance to erythromycin or tetracycline; and natural resistance to gentamicin). A checkerboard technique and a microdilution standardized method were used. All compounds individually tested exhibited antimicrobial activity against E. faecalis, with minimal inhibitory concentrations (MIC) ranging from 30 μg/mL (octyl gallate) to 3,150 μg/mL (gallic acid), although no significant differences were detected among strains to each phenolic compound. Carvacrol in combination with thymol or gallic acid, and gallic acid combined with octyl gallate showed partial synergistic inhibition of all E. faecalis strains. The most effective combinations were thymol+carvacrol and gallic acid+octyl gallate, as the MIC for each of these compounds was reduced by 67 to 75% compared with their respective individual MIC. These results highlight the possibility of using combinations of these phenolic compounds to inhibit the growth of potential virulent or spoilage E. faecalis strains by reducing the total amount of additives used in dairy foods.