Coenzyme Specificity of Enzymes in the Oxidative Pentose Phosphate Pathway of<i>Gluconobacter oxydans</i>

Tonouchi, Naoto; Sugiyama, Masakazu; Yokozeki, Kenzo

doi:10.1271/bbb.67.2648

Cited by 16 publications

(13 citation statements)

References 18 publications

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“…1). The two dehydrogenases of the oxidative PPP were shown to have dual coenzyme specificities (25), but based on the kinetic properties, it was proposed that in vivo glucose-6-phosphate dehydrogenase utilizes NADP ϩ and 6-phosphogluconate dehydrogenase NAD ϩ as cofactors (21). The acetaldehyde dehydrogenase of G. oxydans is NADP dependent (12).…”

Section: Discussionmentioning

confidence: 99%

Mutational Analysis of the Pentose Phosphate and Entner-Doudoroff Pathways in Gluconobacter oxydans Reveals Improved Growth of a Δ edd Δ eda Mutant on Mannitol

Richhardt¹,

Bringer²,

Bott³

2012

Appl Environ Microbiol

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ABSTRACTThe obligatory aerobic acetic acid bacteriumGluconobacter oxydans621H oxidizes sugars and sugar alcohols primarily in the periplasm, and only a small fraction is metabolized in the cytoplasm. The latter can occur either via the Entner-Doudoroff pathway (EDP) or via the pentose phosphate pathway (PPP). The Embden-Meyerhof pathway is nonfunctional, and a cyclic operation of the tricarboxylic acid cycle is prevented by the absence of succinate dehydrogenase. In this work, the cytoplasmic catabolism of fructose formed by oxidation of mannitol was analyzed with a Δgndmutant lacking the oxidative PPP and a ΔeddΔedamutant devoid of the EDP. The growth characteristics of the two mutants under controlled conditions with mannitol as the carbon source and enzyme activities showed that the PPP is the main route for cytoplasmic fructose catabolism, whereas the EDP is dispensable and even unfavorable. The ΔeddΔedamutant (lacking 6-phosphogluconate dehydratase and 2-keto-3-deoxy-6-phosphogluconate aldolase) formed 24% more cell mass than the reference strain. In contrast, deletion ofgnd(6-phosphogluconate dehydrogenase) severely inhibited growth and caused a strong selection pressure for secondary mutations inactivating glucose-6-phosphate dehydrogenase, thus preventing fructose catabolism via the EDP also. These Δgnd zwf* mutants (with a mutation in thezwfgene causing inactivation of the glucose-6-phosphate dehydrogenase) were almost totally disabled in fructose catabolism but still produced about 14% of the carbon dioxide of the reference strain, possibly by catabolizing substrates from the yeast extract. Overexpression ofgndin the reference strain improved biomass formation in a similar manner as deletion ofeddandeda, further confirming the importance of the PPP for cytoplasmic fructose catabolism.

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Section: Discussionmentioning

confidence: 99%

Mutational Analysis of the Pentose Phosphate and Entner-Doudoroff Pathways in Gluconobacter oxydans Reveals Improved Growth of a Δ edd Δ eda Mutant on Mannitol

Richhardt¹,

Bringer²,

Bott³

2012

Appl Environ Microbiol

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“…Again, different dilutions of cell extract were applied. Since 6-phosphogluconate dehydrogenase is an NAD ϩ -preferring enzyme in G. oxydans, 0.1 mM NAD ϩ was used instead of NADP ϩ for activity determination (24,25). The protein concentration of the cell extracts was determined by the method of Bradford (26) by using bovine serum albumin as the standard.…”

Section: Methodsmentioning

confidence: 99%

Combined Fluxomics and Transcriptomics Analysis of Glucose Catabolism via a Partially Cyclic Pentose Phosphate Pathway in Gluconobacter oxydans 621H

Hanke¹,

Nöh²,

Noack³

et al. 2013

Appl Environ Microbiol

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In this study, the distribution and regulation of periplasmic and cytoplasmic carbon fluxes in Gluconobacter oxydans 621H with glucose were studied by 13 C-based metabolic flux analysis ( 13 C-MFA) in combination with transcriptomics and enzyme assays. For 13 C-MFA, cells were cultivated with specifically 13 C-labeled glucose, and intracellular metabolites were analyzed for their labeling pattern by liquid chromatography-mass spectrometry (LC-MS). In growth phase I, 90% of the glucose was oxidized periplasmically to gluconate and partially further oxidized to 2-ketogluconate. Of the glucose taken up by the cells, 9% was phosphorylated to glucose 6-phosphate, whereas 91% was oxidized by cytoplasmic glucose dehydrogenase to gluconate. Additional gluconate was taken up into the cells by transport. Of the cytoplasmic gluconate, 70% was oxidized to 5-ketogluconate and 30% was phosphorylated to 6-phosphogluconate. In growth phase II, 87% of gluconate was oxidized to 2-ketogluconate in the periplasm and 13% was taken up by the cells and almost completely converted to 6-phosphogluconate. Since G. oxydans lacks phosphofructokinase, glucose 6-phosphate can be metabolized only via the oxidative pentose phosphate pathway (PPP) or the Entner-Doudoroff pathway (EDP).13 C-MFA showed that 6-phosphogluconate is catabolized primarily via the oxidative PPP in both phases I and II (62% and 93%) and demonstrated a cyclic carbon flux through the oxidative PPP. The transcriptome comparison revealed an increased expression of PPP genes in growth phase II, which was supported by enzyme activity measurements and correlated with the increased PPP flux in phase II. Moreover, genes possibly related to a general stress response displayed increased expression in growth phase II.

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“…4. The oxidation of G6P to glucuronic acid phosphate is carried out by glucose 6 phosphate dehydrogenase, using NAD as cofactor [34,35].…”

Section: Synthesis Of Cellulose and Acetanmentioning

confidence: 99%

The unified metabolism of Gluconacetobacter entanii in continuous and batch processes

Velasco-Bedrán

López‐Isunza

2007

Process Biochemistry

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Coenzyme Specificity of Enzymes in the Oxidative Pentose Phosphate Pathway ofGluconobacter oxydans

Cited by 16 publications

References 18 publications

Mutational Analysis of the Pentose Phosphate and Entner-Doudoroff Pathways in Gluconobacter oxydans Reveals Improved Growth of a Δ edd Δ eda Mutant on Mannitol

Mutational Analysis of the Pentose Phosphate and Entner-Doudoroff Pathways in Gluconobacter oxydans Reveals Improved Growth of a Δ edd Δ eda Mutant on Mannitol

Combined Fluxomics and Transcriptomics Analysis of Glucose Catabolism via a Partially Cyclic Pentose Phosphate Pathway in Gluconobacter oxydans 621H

The unified metabolism of Gluconacetobacter entanii in continuous and batch processes

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