Search

A heat-stable endogenous CO(2) acceptor has been found in extracts of Rhodospirillum rubrum grown photoheterotrophically on acetate. Evidence is presented which suggests that this factor is propionic acid. Thus, paper and gas chromatographic analyses have indicated that propionic acid is present in boiled extracts prepared from R. rubrum cells. The products of (14)CO(2) fixation obtained with either the boiled extract or propionic acid as the CO(2) acceptor were identical and were identified as methylmalonic acid and succinic acid by paper chromatography. The enzyme which catalyzes the carboxylation of propionyl-coenzyme A (propionyl-CoA carboxylase) was purified from R. rubrum cells grown on acetate and its properties were studied. The enzyme is similar to propionyl-CoA carboxylases isolated from mammalian sources.

Dialyzed extracts of Acetobacter suboxydans ATCC 621 catalyze (14)CO(2) assimilation in the presence of phosphoenolpyruvate and a divalent cation. The formation of (14)C-oxalacetate was demonstrated and found not to be dependent upon the presence of orthophosphate or diphosphonucleotides. Oxalacetate synthesis was stimulated by orthophosphate and inhibited by aspartate. All attempts to demonstrate a reversible carboxylation mechanism have failed. (14)C-aspartate was synthesized when phosphoenolpyruvate, H(14)Co(3) (-), pyridoxal phosphate, and glutamate were added to dialyzed extracts. Chromatographic and spectrophotometric analyses and chemical degradation further demonstrate the presence of a reversible aspartate aminotransferase. The function of oxalacetate synthesis in a bacterium that reportedly lacks an operative tricarboxylic acid cycle is discussed.

The agents of meningopneumonitis (MN) and of trachoma (TR) purified from chick embryo allantoic fluids and yolk sacs, respectively, were shown to produce CO(2) from the C(1) positions of pyruvate and glutamate, but not from the other carbon atoms. The reaction with pyruvate did not require did not require the addition of cofactors, but was stimulated to a small extent by alpha-lipoic acid and, in the case of TR, also by diphosphothiamine, and nicotinamide adenine dinucleotide (NAD). The reaction of MN with glutamate was greatly stimulated by the addition of NAD and pyruvate, and resulted in the accumulation of alanine. The reaction of TR with glutamate was also greatly enhanced by added NAD, but was not affected by added pyruvate. When eight intermediates of the citric acid cycle were added to MN cells incubated with glutamate-C(14), plus NAD and pyruvate, they reduced to varying degrees the evolution of C(14)O(2). It was shown by chromatography that the C(14) label extended to alpha-ketoglutarate and succinate, but not to fumarate and malate. A net gain in adenosine triphosphate could not be demonstrated in MN cells incubated with combined glutamate, pyruvate, oxaloacetate, and various cofactors. These results furnish additional examples of real or apparent gaps in enzyme sequences in Chlamydia.

Extracts of cells of Streptococcus faecalis var. liquefaciens strain 31 incorporated (14)CO(2) into aspartate. Dialyzed extracts produced radioactive oxalacetate in the absence of exogenously added glutamate and pyridoxal-5'-phosphate and produced radioactive aspartate in the presence of these components. Reduced nicotinamide adenine dinucleotide or reduced nicotinamide adenine dinucleotide phosphate could not be substituted for adenosine triphosphate (ATP); phosphoenolpyruvate even in the presence of nucleoside diphosphates could not replace pyruvate plus ATP; propionate plus coenzyme A (CoA) could not replace pyruvate in supporting CO(2) fixation by cell extracts. Fixation by dialyzed cell extracts required pyruvate, ATP, MgSO(4), and was stimulated by biotin, KCl, 2-mercaptoethanol, CoA, and acetyl CoA. Inhibition of fixation occurred when avidin, NaCl, oxalacetate, or aspartate was added to dialyzed extracts. On the basis of the products formed and the effects of substrates and cofactors on the fixation reaction, it was concluded that pyruvate carboxylase is responsible for CO(2) fixation in this microorganism.