The regulatory properties of yeast pyruvate kinase. Effect of pH

Kinderlerer, Julian; Ainsworth, Stanley; Morris, C N; Rhodes, Nicholas P.

doi:10.1042/bj2340699

Cited by 16 publications

(20 citation statements)

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“…The enzyme from yeast has been widely studied and its kinetics are well defined (Haeckel et al, 1968;Kinderlerer et al, 1986;Rhodes et al, 1986). The gene for yeast pyruvate kinase has been cloned and expressed to high levels in the yeast Saccharomyces cerevisiae and this allows easy purification of large amounts of the enzyme (Murcott et al, 1991).…”

Section: Introductionmentioning

confidence: 99%

The cooperative binding of fructose-1,6-bisphosphate to yeast pyruvate kinase.

1992

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The cooperative binding of the allosteric activator fructose‐1,6‐bisphosphate [Fru(1,6)P2] to yeast pyruvate kinase was investigated by equilibrium dialysis and fluorescence quench titration. The results show that yeast pyruvate kinase binds four molecules of Fru(1,6)P2 per tetramer and the observed fluorescence quench follows the binding of the ligand and not the cooperative T to R state transition. Additionally it is shown that the binding of Fru(1,6)P2 to yeast pyruvate kinase is compatible with the model of cooperativity that has been proposed and incorporates an intermediate state, R′, with properties between those of the T and R states.

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Section: Introductionmentioning

confidence: 99%

The cooperative binding of fructose-1,6-bisphosphate to yeast pyruvate kinase.

1992

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“…Its kinetic and regulatory behaviour are well documented (Haeckel et al, 1968;Wieker and Hess, 1971 ;Kinderlerer et al, 1986;Rhodes et al, 1986) and the gene has been sequenced (Burke et al, 1933;McNally et al, 1989) and expressed from multicopy plasmids (Moore et al, 1990). Recombinant liver pyruvate kinase (L-type) has also been expressed in monkey COS cells (Tani et al, 1988), although the levels of expression were low and there was contamination with endogenous pyruvate kinase activity.…”

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confidence: 99%

Purification, characterisation and mutagenesis of highly expressed recombinant yeast pyruvate kinase

Murcott

McNally

Allen

et al. 1991

European Journal of Biochemistry

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Recombinant yeast pyruvate kinase has been purified from a strain of Saccharomyces cerevisiae expressing the enzyme to very high levels. Expression was from a multicopy plasmid under the control of the yeast phosphoglycerate kinase promoter. The gene was expressed in the absence of the genomically encoded pyruvate kinase, using a strain of yeast in which the pyruvate kinase gene has been disrupted by the insertion of the yeast Ura3 gene. The purification procedure minimised proteolytic artefacts and enabled the covenient purification of 15 -20 mg enzyme from 1 1 culture. The purified enzyme was characterised by a high specific activity and by a lack of proteolytic degradation. Two active-site mutants of yeast pyruvate kinase have been produced, expressed and characterised in this system and preliminary results are described.Pyruvate kinase (ATP : pyruvate 2-0-phosphotransferase) is an enzyme of glycolysis that plays a major role in regulating the flux from fructose 1,6-bisphosphate (Fru(l,6)P2) to pyruvate. The enzyme is a homotetramer, with a subunit M , of 55 000 -60000, and catalyses the essentially irreversible reaction,Pyruvate kinase has been extensively studied from a wide range of organisms and much is known about its physical and catalytic properties. It exists as four isoenzymes in mammals designated M1, M2, L and R. The M1 isoenzyme is essentially non-regulated, whereas the remaining isoenzymes (as well as the enzymes from lower organisms) exhibit a sigmoidal kinetic profile with respect to phosphoenolpyruvate concentration, and are allosterically regulated by a large number of nonsubstrate molecules. All forms of the enzyme require both monovalent and bivalent cations for activity. The former is normally potassium whereas a number of bivalent cations will suffice. Two bivalent cations are required/active site, one primarily associated with the nucleotide and the other more closely enzyme associated (see Muirhead, 1987, for a review).The three-dimensional structure of cat muscle M1 isoenzyme has been solved to a resolution of 0.26 nm (Muirhead et al., 1986). Each subunit of cat muscle M1 pyruvate kinase consists of four domains, N, A, B and C. Domain A is an eight-stranded a/B barrel, typical of a number of enzymes (Farber and Petsko, 1990). The active site lies between do-1 Kt, Mg'+

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“…The last and rate-limiting step of glycolysis is catalyzed by pyruvate kinase (PK), converting PEP into pyruvate, and producing one ATP molecule at the same time. The ATP/AMP ratio and intracellular pH have an important effect on PK activity [17] . There are four isoforms of pyruvate kinase: PKL, PKR, PKM1 and PKM2 [18] , among which the expression of PKM2 is upregulated in human tumor cells and cancerassociated fibroblasts [19,20] .…”

Section: The Process Of Glycolysismentioning

confidence: 99%

Anticancer strategies based on the metabolic profile of tumor cells: therapeutic targeting of the Warburg effect

et al. 2016

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Tumor cells rely mainly on glycolysis for energy production even in the presence of sufficient oxygen, a phenomenon termed the Warburg effect, which is the most outstanding characteristic of energy metabolism in cancer cells. This metabolic adaptation is believed to be critical for tumor cell growth and proliferation, and a number of onco-proteins and tumor suppressors, including the PI3K/Akt/mTOR signaling pathway, Myc, hypoxia-inducible factor and p53, are involved in the regulation of this metabolic adaptation. Moreover, glycolytic cancer cells are often invasive and impervious to therapeutic intervention. Thus, altered energy metabolism is now appreciated as a hallmark of cancer and a promising target for cancer treatment. A better understanding of the biology and the regulatory mechanisms of aerobic glycolysis has the potential to facilitate the development of glycolysis-based therapeutic interventions for cancer. In addition, glycolysis inhibition combined with DNA damaging drugs or chemotherapeutic agents may be effective anticancer strategies through weakening cell damage repair capacity and enhancing drug cytotoxicity.

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The regulatory properties of yeast pyruvate kinase. Effect of pH

Cited by 16 publications

References 11 publications

The cooperative binding of fructose-1,6-bisphosphate to yeast pyruvate kinase.

The cooperative binding of fructose-1,6-bisphosphate to yeast pyruvate kinase.

Purification, characterisation and mutagenesis of highly expressed recombinant yeast pyruvate kinase

Anticancer strategies based on the metabolic profile of tumor cells: therapeutic targeting of the Warburg effect

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