Arabinose is metabolized via a phosphoketolase pathway in Clostridium acetobutylicum ATCC 824

Abstract

In this report, a novel zymogram assay and coupled phosphoketolase assay were employed to demonstrate that Clostridium acetobutylicum gene CAC1343 encodes a bi-functional xylulose-5-P/fructose-6-P phosphoketolase (XFP). The specific activity of purified recombinant XFP was 6.9 U/mg on xylulose-5-P and 21 U/mg on fructose-6-P, while the specific activity of XFP in concentrated C. acetobutylicum whole-cell extract was 0.094 and 0.52 U/mg, respectively. Analysis of crude cell extracts indicated that XFP activity was present in cells grown on arabinose but not glucose and quantitative PCR was used to show that CAC1343 mRNA expression was induced 185-fold during growth on arabinose when compared to growth on glucose. HPLC analysis of metabolites revealed that during growth on xylose and glucose more butyrate than acetate was formed with final acetate:butyrate ratios of 0.72 and 0.83, respectively. Growth on arabinose caused a metabolic shift to more oxidized products with a final acetate:butyrate ratio of 1.95. The shift towards more oxidized products is consistent with the presence of an XFP, suggesting that arabinose is metabolized via a phosphoketolase pathway while xylose is probably metabolized via the pentose phosphate pathway.

Erratum: Divergent functional isoforms drive niche specialisation for nutrient acquisition and use in rumen microbiome

“…This is involved in the conversion of carbon compounds in fatty acids (Ellis et al, 2008;Servinsky et al, 2012), and their higher diversity suggests Prevotella has a greater repertoire of enzymes involved in this process and supports previous reports that Prevotella species are among those (including Clostridium species) that play a role in biohydrogenation of dietary polyunsaturated fatty acids to saturated fatty acids (Huws et al, 2011). This would suggest that fatty acids may be a resource for which these organisms compete and for which they have diversified niches.…”

Hierarchy in Pentose Sugar Metabolism in Clostridium acetobutylicum

“…These bacteria ferment polysaccharides from plant detritus, including sugars from cellulose and hemicellulose, into acids and solvents (1)(2)(3)(4)(5). The model organism Clostridium acetobutylicum and other Clostridium species are of considerable commercial interest because of their ability to produce hydrogen gas, acetone, butanol, and ethanol from lignocellulosic components (6)(7)(8).…”

“…Historically, it was assumed that carbons from Xyl and Arab pass from the PPP through glycolysis to supply acidogenesis and solventogenesis. However, two recent studies (5,13) reported that acids and solvents can be produced from these sugars from a glycolysis-independent pathway, the phosphoketolase pathway (PKP) (Fig. 1B).…”

“…The PKP involves cleavage of xylulose-5-phosphate (Xu5P) into glyceraldehyde-3-phosphate (GAP) and acetyl-phosphate (Ac-P) and, to a lesser extent, cleavage of fructose-6-phosphate (F6P) into erythrose-4-phosphate (E4P) and Ac-P (Fig. 1B) (5,13). The metabolite Ac-P is a direct precursor to either acetate or acetyl coenzyme A (acetyl-CoA), which are both involved in clostridial production of acids and solvents (Fig.…”

“…Using 13 C metabolic flux analysis, Liu et al (13) calculated that Xyl catabolic flux through the PKP in C. acetobutylicum increases with increasing Xyl concentration. Furthermore, due to both higher ratios of excreted acetate and a more than 100-fold increase in the mRNA expression of the phosphoketolase enzyme in the presence of Arab, Servinsky et al (5) proposed that the PKP is more significant when the cells are grown with Arab as the C source than with either Gluc or Xyl. However, the significance of this pathway in the metabolism of sugar mixtures was not elucidated.…”