ABSTRACT
Disrupted Ascaris mitochondria formed 2-methylbutyrate (2-MB) and 2-methylvalerate (2-MV) when incubated anaerobically with acetyl CoA, propionyl CoA and NADH. However, when mitochondrial membranes were removed by high speed centrifugation and the mitochondrial soluble fraction was incubated with the same substrates, 2-methylcrotonate (tiglate) and a compound tentatively identified as 2-methyl-2- pentenoate accumulated rather than 2-MB or 2-MV. These data suggest that the terminal reduction of the unsaturated intermediates to the saturated 2-MB and 2-MV was catalyzed by an enzyme system at least partially bound to membranes. This supposition was further supported by the findings that disrupted Ascaris mitochondria also formed 2-MB and lesser amounts of 2-MV when incubated with tiglyl CoA plus NADH, and both soluble and membrane-bound components appear to be involved in this reduction. The possibility that electron transport associated ATP synthesis may be coupled to these reductions remains to be examined.
Subject(s)
Ascaris/metabolism , Fatty Acids, Volatile/biosynthesis , Mitochondria, Muscle/metabolism , Acyl Coenzyme A , Animals , Ascaris/ultrastructure , Butyrates/biosynthesis , Cell-Free System , Crotonates/biosynthesis , Female , Flavin-Adenine Dinucleotide/pharmacology , Hemiterpenes , NAD/metabolism , Oxidation-Reduction , Pentanoic Acids/biosynthesisABSTRACT
Stereospecificity as well as the dependency on reduced pyridine nucleotides of the enzymatic reduction of acetoacetyl CoA and crotonyl CoA in lactating rabbit mammary glands are reported. In the reduction of both acetoacetyl CoA and crotonyl CoA as substrates the tritium from NADP3H was stereospecifically incorporated into the beta-position of n-butyric acid. The reaction of acetoacetyl CoA reduction was much more dependent on NADH while the reduction of crotonyl CoA was rather more dependent on NADPH. There was no difference between the dependencies on NADH and NADPH in the reduction of 2-hexenyl CoA.
Subject(s)
Acetyl Coenzyme A , Coenzyme A/analogs & derivatives , Mammary Glands, Animal/enzymology , Acetyl Coenzyme A/analogs & derivatives , Acetyl Coenzyme A/metabolism , Acyl Coenzyme A , Adenosine Triphosphate/metabolism , Animals , Butyrates/biosynthesis , Chromatography, Paper , Coenzyme A/metabolism , Crotonates/biosynthesis , Fatty Acids/metabolism , Female , Hydroxybutyrates/metabolism , Lactation , NAD/metabolism , NADP/metabolism , Pregnancy , Rabbits , SpectrophotometryABSTRACT
A selection procedure is described which was used to isolate mutants of Pseudomonas putida strain U in the following enzymes of the meta-fission pathway of phenol and cresols: hydrolase, tautomerase, and decarboxylase. Strains deficient in the hydrolase are unable to use either o- or m-cresol as a sole carbon source and were shown to accumulate 2-hydroxy-6-keto-2,4-heptadienoate when incubated in the presence of o- or m-cresol. When 2-hydroxymuconic semialdehyde (plus nicotinamide adenine dinucleotide, oxidized form) was metabolized by phenol-induced extracts of tautomerase-deficient strains, the enol tautomer of 4-oxalocrotonate accumulated and was then converted slowly to the keto tautomer by a nonenzymatic reaction. Phenol-induced extracts of decarboxylase-deficient strains accumulated the keto tautomer of 4-oxalocrotonate from 2-hydroxymuconic semialdehyde (plus nicotinamide adenine dinucleotide, oxidized form). Strains with an inactive decarboxylase are unable to completely metabolize either phenol or p-cresol. Tautomerase-defective strains are unable to grow with p-cresol as the sole carbon source and grow only very slowly on phenol.
