Pyruvate kinase crystallization

Stuart DI, Levine M, Muirhead H, Stammers DK. Crystal structure of cat muscle pyruvate kinase at a resolution of 2.6 AjMol Biol 1979 134 109-142. 

Pyruvate kinase is a key enzyme in the important process of glycolysis, and its product, pyruvate, is passed into the Krebs cycle. As noted in Section 62.1.2.3.1, pyruvate requires two divalent cations and one monovalent cation for activity. These are usually Mg2+ and K+. The crystal structure of the enzyme from cat muscle has been determined to a resolution of 2.6 A.280... 

Pyruvate kinase catalyzes the conversion of phospho-enolpyruvate and ADP to pyruvate and ATP. A proton is taken up in the reaction. The enzyme binds one K+ and two Mg + ions. The 2.9 A crystal structme shows that the K+ interacts directly with the migrating phosphoryl group and indirectly with residues in the active site. Activity with Na+ is about 9% that of K+. Figme 1 shows the geometry of the active site. Oxalyl phosphate is a substrate for pyruvate... 

A. Mattevi, G. Valentini, M. Rizzi, M L. Speranza, M. Bolognesi, and A. Coda. 1995. Crystal structure oiEscherichia coli pyruvate kinase type I Molecular basis of the allosteric transition Structure 3 729-741. (PubMed)... 

Mattevi A, Valentini G, Rizzi M> Speranza ML, Bolog-nesi M, Coda A. Crystal structure of Escherichia coh pyruvate kinase type I molecular basis of the allosteric transition. Structure 1995 3 729-41. 

Mattevi, A., Valentini, G., Rlzzi, M., Speranza, M. L., Bolognesi, M., and Coda, A., 1995. Crystal structure of Escherichia coli pyruvate kinase type 1 Molecular basis of the allosteric transition. Structure 3 729-741. 

Schmidt-Base, K. Buchbinder, J.L. Reed, G.H. Rayment, I. Crystallization and preliminary analysis of enzyme-substrate complexes of pyruvate kinase from rabbit muscle. Proteins Struct. Funct. Genet., 11, 153-157... 

Fothergill-Gillmore, L.A. Rigden, D.J. Michels, P.A. Phillips, S.E. Leish-mania pyruvate kinase the crystal structure reveals the structural basis of its unique regulatory properties. Biochem. Soc. Trans., 28, 186-190 (2000)... 

A reversible reaction catalyzes the conversion of pyruvate to phosphopyruvate, and the enzyme involved is pyruvic kinase. The equilibrium of that reaction is on the side of the formation of ATP. Thus, pyruvate kinase is the enzyme responsible for the conversion of phosphoenolpyruvate to pyruvate. The enzyme has been crystallized from muscle it requires ADP, potassium, and magnesium and is noncompetitively inhibited by some estrogenic steroids. Steroids alter the enzyme s viscosity and electrophoretic properties. From this observation, it was assumed that steroids act by modifying the protein molecule. 

The initial experiments indicated that COi activation and carboxylation might be independent reactions 9S). A COj-dependent reaction was observed in which fluorophosphate was formed from ATP in the absence of propionyl CoA. This led to the tentative assumption that carbonyl phosphate might be active COt. The enzyme catalyzing this reaction was named fluorokinase. Subsequently this enzyme was crystallized and shown to be identical with pyruvic kinase (fiS). 

Coon and co-workers ttt-ltJia) discovered and crystallized an enzyme Itia) named hydroxylamine kinase that degrades ATP in the presence of bicarbonate and hydroxylamine. It was believed that adenylic acid and pyrophosphate were products of this reaction and it was considered that this enzyme participated in the carboxylation reaction in leucine catabolism by activating COi. Active CO was presumed to be adenyl-COj. Later this enzyme was found not to participate in the carboxylation reactions. More recently Kupiecki and Coon lt4b) established that hydroxylamine kinase is very probably identical with pyruvic kinase and fluoro kinase of Tietz and Ochoa (96). Interesting differences were found in the activation of the different enzyme functions by metal ions. Zn, but not Mg++, promoted hydroxylamine kinase activity, whereas Mg, but not Zn promoted iluorokinase and pyruvic kinase activity. 

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