Properties of a pentulose-5-phosphate phosphoketolase from yeasts grown on xylose

Ratledge, Colin; Holdsworth, Jane E.

doi:10.1007/bf00253613

Cited by 19 publications

(6 citation statements)

References 4 publications

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“…These kinetic properties imply that the enzyme prefers to catalyze acetate and ATP synthesis from acetyl phosphate and ADP. Along with the co-transcription of the xfp and ack genes, these features indicate that AcK of M. alcaliphilum 20Z is a member of the phosphoketolase pathway described as a bifid shunt, which is a potential alternative carbon route to the enhancement of acetyl-CoA production in lactic acid bacteria, bifidobacteria, and xylose fermenting yeasts (Ratledge and Holdsworth 1985;Sánchez et al 2010). XFPs are known to be bifunctional enzymes converting hexoses into acetic and lactic acids or directly splitting xylulose-5-phosphate into glyceraldehyde-3-phosphate and acetyl-phosphate bypassing the non-oxidative branch of the pentose phosphate pathway (Sánchez et al 2010).…”

Section: Discussionmentioning

confidence: 97%

Acetate kinase-an enzyme of the postulated phosphoketolase pathway in Methylomicrobium alcaliphilum 20Z

Rozova

Khmelenina

Gavletdinova

et al. 2015

Antonie van Leeuwenhoek

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Recombinant acetate kinase (AcK) was obtained from the aerobic haloalkalitolerant methanotroph Methylomicrobium alcaliphilum 20Z by heterologous expression in Escherichia coli and purification by affinity chromatography. The substrate specificity, the kinetics and oligomeric state of the His6-tagged AcK were determined. The M. alcaliphilum AcK (2 × 45 kDa) catalyzed the reversible phosphorylation of acetate into acetyl phosphate and exhibited a dependence on Mg(2+) or Mn(2+) ions and strong specificity to ATP/ADP. The enzyme showed the maximal activity and high stability at 70 °C. AcK was 20-fold more active in the reaction of acetate synthesis compared to acetate phosphorylation and had a higher affinity to acetyl phosphate (K m 0.11 mM) than to acetate (K m 5.6 mM). The k cat /K m ratios indicated that the enzyme had a remarkably high catalytic efficiency for acetate and ATP formation (k cat/K m = 1.7 × 10(6)) compared to acetate phosphorylation (k cat/K m = 2.5 × 10(3)). The ack gene of M. alcaliphilum 20Z was shown to be co-transcribed with the xfp gene encoding putative phosphoketolase. The Blast analysis revealed the ack and xfp genes in most genomes of the sequenced aerobic methanotrophs, as well as methylotrophic bacteria not growing on methane. The distribution and metabolic role of the postulated phosphoketolase shunted glycolytic pathway in aerobic C1-utilizing bacteria is discussed.

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Section: Discussionmentioning

confidence: 97%

Acetate kinase-an enzyme of the postulated phosphoketolase pathway in Methylomicrobium alcaliphilum 20Z

Rozova

Khmelenina

Gavletdinova

et al. 2015

Antonie van Leeuwenhoek

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“…It is known that xylose enters the pentose phosphate (PP) pathway, but it is not clear how it interacts with the remaining part of the metabolism. Phosphoketolases (EC 4.1.2.9, EC 4.1.2.22) are a class of key enzymes of the phosphoketolase pathway of heterofermentative and facultatively homofermentative lactic acid bacteria, as well as the D-fructose 6-phosphate shunt of bifidobacteria, and xylose fermenting yeast [1] , [2] . The first type of phosphoketolase (PHK, EC 4.1.2.9) catalyses an irreversible thiamine diphosphate dependent phosphorolytic reaction: in the presence of inorganic phosphate, D-xylulose-5-phosphate is cleaved into acetyl phosphate and glyceraldehyde-3-phosphate.…”

Section: Introductionmentioning

confidence: 99%

Systems Analysis Unfolds the Relationship between the Phosphoketolase Pathway and Growth in Aspergillus nidulans

Panagiotou

Andersen

Grotkjær³

et al. 2008

PLoS ONE

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Background Aspergillus nidulans is an important model organism for studies on fundamental eukaryotic cell biology and on industrial processes due to its close relation to A. niger and A. oryzae. Here we identified the gene coding for a novel metabolic pathway in A. nidulans, namely the phosphoketolase pathway, and investigated the role of an increased phosphoketolase activity.Methodology/Principal FindingsOver-expression of the phosphoketolase gene (phk) improved the specific growth rate on xylose, glycerol and ethanol. Transcriptome analysis showed that a total of 1,222 genes were significantly affected by over-expression of the phk, while more than half of the affected genes were carbon source specific. During growth on glucose medium, the transcriptome analysis showed that the response to phk over-expression is targeted to neutralize the effect of the over-expression by regulating the acetate metabolism and initiate a growth dampening response.Conclusions/SignificanceMetabolic flux analysis using 13C-labelled glucose, showed that over-expression of phosphoketolase added flexibility to the central metabolism. Our findings further suggests that A. nidulans is not optimized for growth on xylose, glycerol or ethanol as the sole carbon sources.

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“…In the presence of inorganic phospate this enzyme converts xylulose 5Ј-phosphate (X5P) into glyceraldehyde 3-phosphate and acetylphosphate. Phosphoketolase activity was first detected in heterofermentative lactobacilli (7,9,12,25), but it has also been found in other organisms, such as Acetobacter xylinum (20,23), yeasts (6,21), Thiobacillus novellus (8), and Butyrivibrio fibrisolvens (16), as well as Fibrobacter succinogenes and Fibrobacter intestinalis (17). In bifidobacteria two types of phosphoketolase have been described, a fructose 6-phosphate (F6P)-specific enzyme and an enzyme with specificity for F6P and X5P.…”

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confidence: 99%

Expression of the Xylulose 5-Phosphate Phosphoketolase Gene, xpkA , from Lactobacillus pentosus MD363 Is Induced by Sugars That Are Fermented via the Phosphoketolase Pathway and Is Repressed by Glucose Mediated by CcpA and the Mannose Phosphoenolpyruvate Phosphotransferase System

Posthuma

Bader

Engelmann

et al. 2002

Appl Environ Microbiol

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Purification of xylulose 5-phosphate phosphoketolase (XpkA), the central enzyme of the phosphoketolase pathway (PKP) in lactic acid bacteria, and cloning and sequence analysis of the encoding gene, xpkA, from Lactobacillus pentosus MD363 are described. xpkA encodes a 788-amino-acid protein with a calculated mass of 88,705 Da. Expression of xpkA in Escherichia coli led to an increase in XpkA activity, while an xpkA knockout mutant of L. pentosus lost XpkA activity and was not able to grow on energy sources that are fermented via the PKP, indicating that xpkA encodes an enzyme with phosphoketolase activity. A database search revealed that there are high levels of similarity between XpkA and a phosphoketolase from Bifidobacterium lactis and between XpkA and a (putative) protein present in a number of evolutionarily distantly related organisms (up to 54% identical residues). Expression of xpkA in L. pentosus was induced by sugars that are fermented via the PKP and was repressed by glucose mediated by carbon catabolite protein A (CcpA) and by the mannose phosphoenolpyruvate phosphotransferase system. Most of the residues involved in correct binding of the cofactor thiamine pyrophosphate (TPP) that are conserved in transketolase, pyruvate decarboxylase, and pyruvate oxidase were also conserved at a similar position in XpkA, implying that there is a similar TPP-binding fold in XpkA.Lactic acid bacteria (LAB) are capable of generating energy by homo-or heterofermentative degradation of sugars. During anaerobic growth of obligately homofermentative LAB in the presence of excess substrate, energy sources like glucose are converted into pyruvate via the Embden-Meyerhoff-Parnas pathway, and the pyruvate is further metabolized to lactate. During heterofermentative degradation sugars are metabolized via the phosphoketolase pathway (PKP), which results in equimolar amounts of CO 2 , lactate, and acetate-ethanol. Heterolactic LAB can be divided into obligately heterofermentative species, in which both hexoses and pentoses are fermented via the PKP, and facultatively heterofermentative organisms, which degrade hexoses via the Embden-Meyerhoff-Parnas pathway and pentoses via the PKP.Xylulose 5-phosphate phosphoketolase is the central enzyme of the PKP. In the presence of inorganic phospate this enzyme converts xylulose 5Ј-phosphate (X5P) into glyceraldehyde 3-phosphate and acetylphosphate. Phosphoketolase activity was first detected in heterofermentative lactobacilli (7,9,12,25), but it has also been found in other organisms, such as Acetobacter xylinum (20, 23), yeasts (6, 21), Thiobacillus novel-

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Properties of a pentulose-5-phosphate phosphoketolase from yeasts grown on xylose

Cited by 19 publications

References 4 publications

Acetate kinase-an enzyme of the postulated phosphoketolase pathway in Methylomicrobium alcaliphilum 20Z

Acetate kinase-an enzyme of the postulated phosphoketolase pathway in Methylomicrobium alcaliphilum 20Z

Systems Analysis Unfolds the Relationship between the Phosphoketolase Pathway and Growth in Aspergillus nidulans

Expression of the Xylulose 5-Phosphate Phosphoketolase Gene, xpkA , from Lactobacillus pentosus MD363 Is Induced by Sugars That Are Fermented via the Phosphoketolase Pathway and Is Repressed by Glucose Mediated by CcpA and the Mannose Phosphoenolpyruvate Phosphotransferase System

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