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Volume 156, Issue 6

Metabolic flux analysis of wild-type and mutants deficient in pyruvate-dissimilating enzymes during the fermentative metabolism of glucuronate Free

Abstract

The fermentative metabolism of -glucuronic acid (glucuronate) in was investigated with emphasis on the dissimilation of pyruvate via pyruvate formate-lyase (PFL) and pyruvate dehydrogenase (PDH). and metabolic flux analysis (MFA) revealed that PFL and PDH share the dissimilation of pyruvate in wild-type MG1655. Surprisingly, it was found that PDH supports fermentative growth on glucuronate in the absence of PFL. The PDH-deficient strain (Pdh−) exhibited a slower transition into the exponential phase and a decrease in specific rates of growth and glucuronate utilization. Moreover, a significant redistribution of metabolic fluxes was found in PDH- and PFL-deficient strains. Since no role had been proposed for PDH in the fermentative metabolism of , the metabolic differences between MG1655 and Pdh− were further investigated. An increase in the oxidative pentose phosphate pathway (ox-PPP) flux was observed in response to PDH deficiency. A comparison of the ox-PPP and PDH pathways led to the hypothesis that the role of PDH is the supply of reducing equivalents. The finding that a PDH deficiency lowers the NADH : NAD ratio supported the proposed role of PDH. Moreover, the NADH : NAD ratio in a strain deficient in both PDH and the ox-PPP (Pdh−Zwf−) was even lower than that observed for Pdh−. Strain Pdh−Zwf− also exhibited a slower transition into the exponential phase and a lower growth rate than Pdh−. Finally, a transhydrogenase-deficient strain grew more slowly than wild-type but did not show the slower transition into the exponential phase characteristic of Pdh− mutants. It is proposed that PDH fulfils two metabolic functions. First, by creating the appropriate internal redox state (i.e. appropriate NADH : NAD ratio), PDH ensures the functioning of the glucuronate utilization pathway. Secondly, the NADH generated by PDH can be converted to NADPH by the action of transhydrogenases, thus serving as biosynthetic reducing power in the synthesis of building blocks and macromolecules.

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Keyword(s): AcCoA, acetyl-CoA , ACK, acetate kinase , MFA, metabolic flux analysis , non-ox-PPP, non-oxidative pentose phosphate pathway , ox-PPP, oxidative pentose phosphate pathway , PDH, pyruvate dehydrogenase , PFL, pyruvate formate-lyase , PP, pentose phosphate , PPP, pentose phosphate pathway , PTA, phosphotransacetylase and TCA cycle, tricarboxylic acid cycle
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2010-06-01
2024-04-25
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Metabolic flux analysis of wild-type Escherichia coli and mutants deficient in pyruvate-dissimilating enzymes during the fermentative metabolism of glucuronate
Microbiology 156, 1860 (2010); https://doi.org/10.1099/mic.0.036251-0
/content/journal/micro/10.1099/mic.0.036251-0
/content/journal/micro/10.1099/mic.0.036251-0
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