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Volume 147, Issue 8

The methylcitric acid pathway in : new genes identified involved in propionate metabolism

The GenBank accession numbers for the nucleotide sequences of the gene cluster are AF325554 and AF331923.

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Abstract

From HF39 null-allele mutants were created by Tn mutagenesis and by homologous recombination which were impaired in growth on propionic acid and levulinic acid. From the molecular, physiological and enzymic analysis of these mutants it was concluded that in this bacterium propionic acid is metabolized via the methylcitric acid pathway. The genes encoding enzymes of this pathway are organized in a cluster in the order , , , , ORF5 and , with transcribed divergently from the other genes. (i) encodes a 2-methylcitric acid synthase (42720 Da) as shown by the measurement of the respective enzyme activity, complementation of a mutant of serovar Typhimurium and high sequence similarity. (ii) For the translational product of the function of a 2-methyl--aconitic acid hydratase (94726 Da) is proposed. This protein and also the ORF5 translational product are essential for growth on propionic acid, as revealed by the propionic-acid-negative phenotype of Tn-insertion mutants, and are required for the conversion of 2-methylcitric acid into 2-methylisocitric acid as shown by the accumulation of the latter, which could be purified as its calcium salt from the supernatants of these mutants. In contrast, inactivation of did not block the ability of the cell to use propionic acid as carbon and energy source, as shown by the propionic acid phenotype of a null-allele mutant. It is therefore unlikely that from encodes a 2-methyl--aconitic acid dehydratase as proposed recently for the homologous gene from . (iii) The translational product of encodes 2-methylisocitric acid lyase (32314 Da) as revealed by measurement of the respective enzyme activity and by demonstrating accumulation of methylisocitric acid in the supernatant of a null-allele mutant. (iv) The expression of and probably also of the other enzymes is regulated and is induced during cultivation on propionic acid or levulinic acid. The putative translational product of (70895 Da) exhibited high similarities to PrpR of and , and might represent a transcriptional activator of the sigma-54 family involved in the regulation of the other genes. Since the locus of was very different from those of and , an extensive comparison of loci available from databases and literature was done, revealing two different classes of loci.

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  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): 2-methylcitric acid cycle , DIG, digoxigenin , methylcitric acid synthase , propionic acid catabolism , propionyl-CoA and Ralstonia eutropha
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2001-08-01
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The methylcitric acid pathway in Ralstonia eutropha: new genes identified involved in propionate metabolismThe GenBank accession numbers for the nucleotide sequences of the prp gene cluster are AF325554 and AF331923.
Microbiology 147, 2203 (2001); https://doi.org/10.1099/00221287-147-8-2203
/content/journal/micro/10.1099/00221287-147-8-2203
/content/journal/micro/10.1099/00221287-147-8-2203
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