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Query: EC:4.1.1.32 (phosphoenolpyruvate carboxykinase)
 4,204 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Kinetic studies were done to obtain a quantitative estimation of the synergistic interactions that occur between phosphoenolpyruvate carboxylase (orthophosphate:oxaloacetate carboxylase (phosphorylating) E.C. 4.1.1.31) from Escherichia coli K12 and various combinations of its primary substrate, P-enolpyruvate, and the activators acetylcoenzyme A, CDP, GTP, and fructose 1,6-bisphosphate. The analysis involves the evaluation of apparent K values, KS for P-enolpyru;ate and KA for activators, as a function of the concentration of P-enolpyruvate or another activator in the case of KA determinations. Methods are presented which allow the determination of dissociation constants for P-enolpyruvate and activators from binary, ternary, and quaternary complexes of enzyme with substrates or activators, or both. It was observed that synergistic activation occurs with acetyl coenzyme A and all of the coactivators but not with fructose 1,6-bisphosphate and the other co-activators. The enhancement of binding from the binary enzyme substrate (or activator) complex to the ternary or quaternary complexes is in the range of 100-fold. The dissociation constants for P-enolpyruvate, acetyl coenzyme A, CDP, and fructose 1,6-bisphosphate are the same from any active enzyme species. Synergistic activation by multiple activators reflects the ability of co-activators to shift the equilibrium concentrations of active enzyme species away from the inactive forms via a modified "cascade" scheme, thus decreasing the probability that dissociation of any one activator will yield an inactive enzyme species.
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PMID:Escherichia coli phosphoenolpyruvate carboxylase. Studies on the mechanism of synergistic activation by nucleotides. 698 77

Cytosolic pyruvate kinase (PKc) from Brassica napus suspension cells was purified 201-fold to electrophoretic homogeneity and a final specific activity of 51 micromol phosphoenolpyruvate utilized per min per mg protein. SDS/PAGE and gel filtration analyses of the final preparation indicated that this PKc is a 220-kDa homotetramer composed of 56-kDa subunits. The enzyme was relatively heat-stable and displayed a broad pH optimum of pH 6.8. PKc activity was absolutely dependent upon the simultaneous presence of a bivalent and univalent cation, with Mg2+ and K+ fulfilling this requirement. Hyperbolic saturation kinetics were observed for phosphoenolpyruvate, ADP, Mg2+ and K+ (apparent Km values = 0.12, 0.075, 0.21 and 0.48 mM, respectively). Although the enzyme utilized UDP, CDP and IDP as alternative nucleotides, ADP was the preferred substrate. L-Glutamate, oxalate, and the flavonoids rutin and quercetin were the most effective inhibitors (I50 values = 4, 0.3, 0.07, and 0.10 mM, respectively). L-Aspartate functioned as an activator (Ka = 0.31 mM) by causing a 40% increase in Vmax while completely reversing the inhibition of PKc by L-glutamate. Reciprocal control by L-aspartate and L-glutamate is specific for these amino acids and provides a rationale for the in vivo activation of PKc that occurs during periods of enhanced NH +4-assimilation. Allosteric features of B. napus PKc are compared with those of B. napus phosphoenolpyruvate carboxylase. A model is presented that highlights the pivotal role of L-aspartate and L-glutamate in the coordinate regulation of these key phosphoenolpyruvate utilizing cytosolic enzymes.
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PMID:Purification and characterization of cytosolic pyruvate kinase from Brassica napus (rapeseed) suspension cell cultures: implications for the integration of glycolysis with nitrogen assimilation. 1088 Sep 71

Cytosolic pyruvate kinase (PK(c)) from ripened banana (Musa cavendishii L.) fruits has been purified 543-fold to electrophoretic homogeneity and a final specific activity of 59.7 micromol of pyruvate produced/min per mg of protein. SDS/PAGE and gel-filtration FPLC of the final preparation indicated that this enzyme exists as a 240 kDa homotetramer composed of subunits of 57 kDa. Although the enzyme displayed a pH optimum of 6.9, optimal efficiency in substrate utilization [in terms of V(max)/K(m) for phosphoenolpyruvate (PEP) or ADP] was equivalent at pH 6.9 and 7.5. PK(c) activity was absolutely dependent upon the presence of a bivalent and a univalent cation, with Mg(2+) and K(+) respectively fulfilling this requirement. Hyperbolic saturation kinetics were observed for the binding of PEP, ADP, Mg(2+) and K(+) (K(m) values of 0.098, 0.12, 0.27 and 0.91 mM respectively). Although the enzyme utilized UDP, IDP, GDP and CDP as alternative nucleotides, ADP was the preferred substrate. L-Glutamate and MgATP were the most effective inhibitors, whereas L-aspartate functioned as an activator by reversing the inhibition of PK(c) by L-glutamate. The allosteric features of banana PK(c) are compared with those of banana PEP carboxylase [Law and Plaxton (1995) Biochem. J. 307, 807-816]. A model is presented which highlights the roles of cytosolic pH, MgATP, L-glutamate and L-aspartate in the co-ordinate control of the PEP branchpoint in ripening bananas.
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PMID:Purification and characterization of cytosolic pyruvate kinase from banana fruit. 1110 98