Purification and properties of pyruvate kinase from Thermoplasma acidophilum

Potter, Simon; Fothergill‐Gilmore, Linda A.

doi:10.1016/0378-1097(92)90636-3

Cited by 8 publications

(7 citation statements)

References 13 publications

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“…This glutamate is also present in the K ϩ -dependent Tpl. acidophilum PK (233). Furthermore, most of the archaeal PKs exhibited positive cooperativity toward PEP and ADP, and the Tpt.…”

Section: Phosphoenolpyruvate Conversion To Pyruvatementioning

confidence: 97%

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Carbohydrate Metabolism in Archaea: Current Insights into Unusual Enzymes and Pathways and Their Regulation

Bräsen

Esser

Rauch

et al. 2014

Microbiol Mol Biol Rev

280

294

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SUMMARY The metabolism of Archaea , the third domain of life, resembles in its complexity those of Bacteria and lower Eukarya . However, this metabolic complexity in Archaea is accompanied by the absence of many “classical” pathways, particularly in central carbohydrate metabolism. Instead, Archaea are characterized by the presence of unique, modified variants of classical pathways such as the Embden-Meyerhof-Parnas (EMP) pathway and the Entner-Doudoroff (ED) pathway. The pentose phosphate pathway is only partly present (if at all), and pentose degradation also significantly differs from that known for bacterial model organisms. These modifications are accompanied by the invention of “new,” unusual enzymes which cause fundamental consequences for the underlying regulatory principles, and classical allosteric regulation sites well established in Bacteria and Eukarya are lost. The aim of this review is to present the current understanding of central carbohydrate metabolic pathways and their regulation in Archaea . In order to give an overview of their complexity, pathway modifications are discussed with respect to unusual archaeal biocatalysts, their structural and mechanistic characteristics, and their regulatory properties in comparison to their classic counterparts from Bacteria and Eukarya . Furthermore, an overview focusing on hexose metabolic, i.e., glycolytic as well as gluconeogenic, pathways identified in archaeal model organisms is given. Their energy gain is discussed, and new insights into different levels of regulation that have been observed so far, including the transcript and protein levels (e.g., gene regulation, known transcription regulators, and posttranslational modification via reversible protein phosphorylation), are presented.

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“…This glutamate is also present in the K ϩ -dependent Tpl. acidophilum PK (233). Furthermore, most of the archaeal PKs exhibited positive cooperativity toward PEP and ADP, and the Tpt.…”

Section: Phosphoenolpyruvate Conversion To Pyruvatementioning

confidence: 97%

“…pernix, and Pyb. aerophilum (232)(233)(234)(235). As for their bacterial and eukaryotic counterparts, the archaeal PKs represent 200-kDa homotetrameric proteins composed of 50-kDa subunits and require divalent metal ions (Mg 2ϩ and Mn 2ϩ ) for activity.…”

Section: Phosphoenolpyruvate Conversion To Pyruvatementioning

confidence: 99%

Carbohydrate Metabolism in Archaea: Current Insights into Unusual Enzymes and Pathways and Their Regulation

Bräsen

Esser

Rauch

et al. 2014

Microbiol Mol Biol Rev

280

294

View full text Add to dashboard Cite

show abstract

“…In contrast to bacterial and eucaryal enzymes, little is known about the structure and function of archaeal PKs. Up to now only the enzymic properties of the PK of Thermoplasma acidophilum could be determined (34). The enzyme represents a homomeric tetramer with a molecular mass of 250 kDa and displays cooperative substrate binding; its substrate affinity is increased by AMP.…”

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confidence: 99%

Pyruvate Kinase of the Hyperthermophilic Crenarchaeote Thermoproteus tenax : Physiological Role and Phylogenetic Aspects

et al. 2000

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Pyruvate kinase (PK; EC 2.7.1.40) of Thermoproteus tenax was purified to homogeneity, and its coding gene was cloned and expressed in Escherichia coli. It represents a homomeric tetramer with a molecular mass of 49 kDa per subunit. PK exhibits positive binding cooperativity with respect to phosphoenolpyruvate and metal ions such as Mg 2؉ and Mn 2؉ . Heterotropic effects, as commonly found for PKs from bacterial and eucaryal sources, could not be detected. The enzyme does not depend on K ؉ ions. Heterotrophically grown cells exhibit specific activity of PK four times higher than autotrophically grown cells. Since the mRNA level of the PK coding gene is also accordingly higher in heterotrophic cells, we conclude that the PK activity is adjusted to growth conditions mainly on the transcript level. The enzymic properties of the PK and the regulation of its expression are discussed with respect to the physiological framework given by the T. tenax-specific variant of the Embden-Meyerhof-Parnas pathway. T. tenax PK shows moderate overall sequence similarity (25 to 40% identity) to its bacterial and eucaryal pendants. Phylogenetic analyses of the known PK sequences result in a dichotomic tree topology that divides the enzymes into two major PK clusters, probably diverged by an early gene duplication event. The phylogenetic divergence is paralleled by a striking phenotypic differentiation of PKs: PKs of cluster I, which occur in eucaryal cytoplasm, some gamma proteobacteria, and low-GC grampositive bacteria, are only active in the presence of fructose-1,6-bisphosphate or other phosphorylated sugars, whereas PKs of cluster II, found in various bacterial phyla, plastids, and in Archaea, show activity without effectors but are commonly regulated by the energy charge of the cell.As shown by several investigations, Archaea use the Embden-Meyerhof-Parnas (EMP) pathway for carbon metabolism, and in some archaeal species (the genera Pyrococcus, Thermococcus, Desulfurococcus, and Thermoproteus) the catabolic EMP pathway represents the main route for glucose degradation (37, 40). So far, in thermophilic Archaea only variants of the classical EMP pathway have been found. Common to these variants is the irreversible oxidation step transforming glyceraldehyde-3-phosphate directly to 3-phosphoglycerate by a ferredoxin-dependent glyceraldehyde-3-phosphate oxidoreductase (GAP:FdOR) found in Pyrococcus, Thermococcus, and Desulfurococcus strains (37, 43) or by a nonphosphorylating NAD ϩ -dependent glyceraldehyde-3-phosphate dehydrogenase (GAPN) found in Thermoproteus tenax (4,15,38). These enzymes substitute for the combined action of conventional phosphorylating-but reversible-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and 3-phosphoglycerate kinase, which, however, allows ATP formation.We chose T. tenax, a hyperthermophilic, facultatively heterotrophic member of the kingdom Crenarchaeota, as an object for regulatory studies on archaeal carbohydrate metabolism. T. tenax is able to grow chemolithoautotrophically on CO 2 , H 2 and S ...

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“…Three enzymes appear to be involved in the crucial PEP/pyruvate inter‐conversion in M. acetivorans . In the glycolytic direction, PYK of M. acetivorans resembles that of Thermoplasma acidophilum because the enzyme activity was fully dependent on K + and NH 4 + . Also, PYK from M. acetivorans showed positive (AMP and alanine) and negative (ATP) modulators.…”

Section: Discussionmentioning

confidence: 97%

The nutritional status of Methanosarcina acetivorans regulates glycogen metabolism and gluconeogenesis and glycolysis fluxes

Santiago-Martínez¹,

Encalada²,

Lira-Silva³

et al. 2016

The FEBS Journal

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Gluconeogenesis is an essential pathway in methanogens because they are unable to use exogenous hexoses as carbon source for cell growth. With the aim of understanding the regulatory mechanisms of central carbon metabolism in Methanosarcina acetivorans, the present study investigated gene expression, the activities and metabolic regulation of key enzymes, metabolite contents and fluxes of gluconeogenesis, as well as glycolysis and glycogen synthesis/degradation pathways. Cells were grown with methanol as a carbon source. Key enzymes were kinetically characterized at physiological pH/temperature. Active consumption of methanol during exponential cell growth correlated with significant methanogenesis, gluconeogenic flux and steady glycogen synthesis. After methanol exhaustion, cells reached the stationary growth phase, which correlated with the rise in glycogen consumption and glycolytic flux, decreased methanogenesis, negligible acetate production and an absence of gluconeogenesis. Elevated activities of carbon monoxide dehydrogenase/acetyl-CoA synthetase complex and pyruvate: ferredoxin oxidoreductase suggested the generation of acetyl-CoA and pyruvate for glycogen synthesis. In the early stationary growth phase, the transcript contents and activities of pyruvate phosphate dikinase, fructose 1,6-bisphosphatase and glycogen synthase decreased, whereas those of glycogen phosphorylase, ADP-phosphofructokinase and pyruvate kinase increased. Therefore, glycogen and gluconeogenic metabolites were synthesized when an external carbon source was provided. Once such a carbon source became depleted, glycolysis and methanogenesis fed by glycogen degradation provided the ATP supply. Weak inhibition of key enzymes by metabolites suggested that the pathways evaluated were mainly transcriptionally regulated. Because glycogen metabolism and glycolysis/gluconeogenesis are not present in all methanogens, the overall data suggest that glycogen storage might represent an environmental advantage for methanosarcinales when carbon sources are scarce. Also, the understanding of the central carbohydrate metabolism in methanosarcinales may help to optimize methane production.Abbreviations 1,3BPG, 1,3-bisphosphoglycerate; 2PG, 2-phosphoglycerate; 3PG, 3-phosphoglycerate; AcK, acetate kinase; ALDO, fructose 1,6-bisphosphate aldolase; CODH/ACS, carbon monoxide dehydrogenase/acetyl-CoA synthetase complex; DHAP, dihydroxyacetone phosphate; Eh-, recombinant enzyme of Entamoeba histolytica; EMP, Embden-Meyerhoff-Parnas; ENO, enolase; Erythro4P, erythrose 4-phosphate; Fru1,6BP, fructose 1,6-bisphosphate; Fru6P, fructose 6-phosphate; FruBPase, fructose 1,6-bisphosphatase; G3P, glyceraldehyde 3-phosphate; G6PDH, glucose 6-phosphate dehydrogenase; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; GAPN, nonphosphorylating GAPDH; GAPOR, glyceraldehyde 3-phosphate: ferredoxin oxidoreductase; Glc1P, glucose 1-phosphate; Glc6P, glucose

show abstract

Purification and properties of pyruvate kinase from Thermoplasma acidophilum

Cited by 8 publications

References 13 publications

Carbohydrate Metabolism in Archaea: Current Insights into Unusual Enzymes and Pathways and Their Regulation

Carbohydrate Metabolism in Archaea: Current Insights into Unusual Enzymes and Pathways and Their Regulation

Pyruvate Kinase of the Hyperthermophilic Crenarchaeote Thermoproteus tenax : Physiological Role and Phylogenetic Aspects

The nutritional status of Methanosarcina acetivorans regulates glycogen metabolism and gluconeogenesis and glycolysis fluxes

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