Identification and molecular characterization of the aco genes encoding the Pelobacter carbinolicus acetoin dehydrogenase enzyme system

Abstract

Use of oligonucleotide probes, which were deduced from the N-terminal sequences of the purified enzyme components, identified the structural genes for the alpha and beta subunits of E1 (acetoin:2,6-dichlorophenolindophenol oxidoreductase), E2 (dihydrolipoamide acetyltransferase), and E3 (dihydrolipoamide dehydrogenase) of the Pelobacter carbinolicus acetoin dehydrogenase enzyme system, which were designated acoA, acoB, acoC, and acoL, respectively. The nucleotide sequences of acoA (979 bp), acoB (1,014 bp), acoC (1,353 bp), and acoL (1,413 bp) as well as of acoS (933 bp), which encodes a protein with an M(r) of 34,421 exhibiting 64.7% amino acid identity to the Escherichia coli lipA gene product, were determined. These genes are clustered on a 6.1-kbp region. Heterologous expression of acoA, acoB, acoC, acoL, and acoS in E. coli was demonstrated. The amino acid sequences deduced from acoA, acoB, acoC, and acoL for E1 alpha (M(r), 34,854), E1 beta (M(r), 36,184), E2 (M(r), 47,281), and E3 (M(r), 49,394) exhibited striking similarities to the amino acid sequences of the components of the Alcaligenes eutrophus acetoin-cleaving system. Homologies of up to 48.7% amino acid identity to the primary structures of the enzyme components of various 2-oxo acid dehydrogenase complexes also were found. In addition, the respective genes of the 2-oxo acid dehydrogenase complexes and of the acetoin dehydrogenase enzyme system were organized very similarly, indicating a close relationship of the P. carbinolicus acetoin dehydrogenase enzyme system to 2-oxo acid dehydrogenase complexes.

Acetoin catabolic system of Klebsiella pneumoniae CG43: sequence, expression, and organization of the aco operon

“…A significant homology was observed among the acoABCD operon of K pneumoniae CG43, the acoXABC operon of A. eutrophus (32), and the aco genes of P. carbinolicus (26). The very high degree of similarity in the deduced amino acid sequences corresponding to the two subunits of El (Fig.…”

The Metabolic Regulation of Sporulation and Parasporal Crystal Formation in Bacillus thuringiensis Revealed by Transcriptomics and Proteomics

“…Meanwhile, the proteins AcuA/B/C were detected by iTRAQ and exhibited slight increases during the stationary growth phase. More importantly, the acoABCL operon encoding the acetoin dehydrogenase complex (47) was strongly up-regulated at both the transcriptional and translational levels at 13 h, likely to cleave reimported acetoin into acetaldehyde and acetyl-CoA (48). Among the three homologues (CH1215, CH2825 and CH3498) of dhaS (aldehyde dehydrogenase), CH1215 and CH3498 were obviously increased at the transcriptional level during the stationary growth phase, furthermore, the protein abundance of CH3498 was also increased by 3.1-fold at both 9 h and 13 h. Therefore, other than being directly converted into acetyl-CoA by ProA (bifunctional acetaldehyde-CoA/alcohol dehydrogenase), acetaldehyde would be sequentially transformed into acetyl-CoA by DhaS, AckA (acetate kinase), and EutD (phosphotransacetylase) (supplemental Fig.…”

Lipoic Acid Metabolism in Microbial Pathogens

“…The presence of an E3 subunit encoded in this cluster varies by species (256), and in cases where it is absent, a common E3 is presumably shared between the AoDH and the ␣-ketoacid dehydrogenases. Interestingly, in P. carbinolicus an additional gene that encodes lipoate synthase is sandwiched between the genes encoding the AoDH E2 and E3 (163), possibly linking expression of lipoylated metabolic complexes and expression of lipoylating enzymes.…”