2022
DOI: 10.1128/aem.01857-21
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Functional Analysis of H + -Pumping Membrane-Bound Pyrophosphatase, ADP-Glucose Synthase, and Pyruvate Phosphate Dikinase as Pyrophosphate Sources in Clostridium thermocellum

Abstract: The atypical glycolysis of Clostridium thermocellum is characterized by the use of pyrophosphate (PP i ) as phosphoryl donor for phosphofructokinase (Pfk) and pyruvate phosphate dikinase (Ppdk) reactions. Previously, biosynthetic PP i was calculated to be stoichiometrically insufficient to drive glycolysis. This study investigates the role of a H + -pumping membrane-bound pyrophosphatase, glycogen cycling, a predict… Show more

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Cited by 14 publications
(21 citation statements)
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“…The sulfur-limited medium allows for a short period of initial growth, where the remaining sulfur sources (mainly cysteine) are consumed, followed by a period of growth-arrest where cells continuously ferment cellobiose to mainly acetate and lactate, and in the absence of added ethanol, to ethanol as main fermentation products (Additional le 4). Given that in C. thermocellum carbon balances of cultivations are known to close poorly (up to 60 -90%) (14,22,(43)(44)(45), using the measured fermentation products as a proxy for the fermentative capacity of growth-arrested cells would result in a signi cant underestimation of this value. Therefore, the fermentative capacity was determined from the change of the fermented cellobiose concentration, which is calculated by assuming that all cellobiose consumed by growth-arrested cells minus the cellobiose that is hydrolyzed to glucose is fermented.…”
Section: Resultsmentioning
confidence: 99%
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“…The sulfur-limited medium allows for a short period of initial growth, where the remaining sulfur sources (mainly cysteine) are consumed, followed by a period of growth-arrest where cells continuously ferment cellobiose to mainly acetate and lactate, and in the absence of added ethanol, to ethanol as main fermentation products (Additional le 4). Given that in C. thermocellum carbon balances of cultivations are known to close poorly (up to 60 -90%) (14,22,(43)(44)(45), using the measured fermentation products as a proxy for the fermentative capacity of growth-arrested cells would result in a signi cant underestimation of this value. Therefore, the fermentative capacity was determined from the change of the fermented cellobiose concentration, which is calculated by assuming that all cellobiose consumed by growth-arrested cells minus the cellobiose that is hydrolyzed to glucose is fermented.…”
Section: Resultsmentioning
confidence: 99%
“…All plasmids used in this study are listed in Table 3. Deletion and integration plasmids were constructed as described previously (43). DNA fragments were PCR ampli ed using pDGO145 or genomic DNA of C. thermocellum DSM1313 as template.…”
Section: Methodsmentioning
confidence: 99%
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“…Glycogen determination. The sampling and assay of glycogen content was performed as previously described (25). Briefly, 1 mL of sample was mixed with 5 mL of ice-cold methanol (-80°C) and centrifuged (10,000 x g, 10 min, -10°C).…”
Section: Methodsmentioning
confidence: 99%