This enzyme belongs to the family of isomerases, specifically the phosphotransferases (phosphomutases), which transfer phosphate groups within a molecule. The systematic name of this enzyme class is phosphoenolpyruvate 2,3-phosphonomutase. Other names in common use include phosphoenolpyruvate-phosphonopyruvate phosphomutase, PEP phosphomutase, phosphoenolpyruvate phosphomutase, PEPPM, and PEP phosphomutase. This enzyme participates in aminophosphonate metabolism.
Phosphoenolpyruvate mutase was discovered in 1988.[1][2]
Structural studies
As of late 2007, 6 structures have been solved for this class of enzymes, all by the Herzberg group at the University of Maryland using PEPPM from the blue mussel, Mytilus edulis. The first structure (PDB accession code 1PYM) was solved in 1999 and featured a magnesium oxalate inhibitor.[3] This structure identified the enzyme as consisting of identical beta barrel subunits (exhibiting the TIM barrel fold, which consists of eight parallel beta strands). Dimerization was observed in which a helix from each subunit interacts with the other subunit's barrel; the authors called this feature "helix swapping." The dimers can dimerize as well to form a homotetrameric enzyme. A double phosphoryl transfer mechanism was proposed on the basis of this study: this would involve breakage of PEP's phosphorus-oxygen bond to form a phosphoenzyme intermediate, followed by transfer of the phosphoryl group from the enzyme to carbon-3, forming PPR.
However, more recently, a structure with a sulfopyruvate inhibitor, which is a closer substrate analogue, was solved (1M1B);[4] this study supported instead a dissociative mechanism. A notable feature of these structures was the shielding of the active site from solvent; it was proposed that a significant conformational change takes place on binding to allow this, moving the protein from an "open" to a "closed" state, and this was supported by several crystal structures in the open state.[5] Three of these were of the wild type: the apoenzyme in 1S2T, the enzyme plus its magnesium ion cofactor in 1S2V, and the enzyme at high ionic strength in 1S2W. A mutant (D58A, in one of the active-site loops) was crystallized as an apoenzyme also (1S2U). From these structures, an active-site "gating" loop (residues 115-133) that shields the substrate from solvent in the closed conformation was identified.
The two conformations, taken from the crystal structures 1M1B (closed) and 1S2T (open), are docked into each other in the images below; they differ negligibly except in the gating loop, which is colored purple for the closed conformation and blue for the open conformation. In the active-site closeup (left), several sidechains (cyan) that have been identified as important in catalysis are included as well; the overview (right) illustrates the distinctive helix-swapping fold. The images are still shots from ribbonkinemages. Both of these structures were crystallized as dimers. In chain A (used for the active-site closeup), helices are red while loops (other than the gating loop) are white and beta strands are green; in chain B, helices are yellow, beta strands are olive, and loops are gray; these colors are the same for the closed and open structures. Magnesium ions are gray and the sulfopyruvate ligands are pink; both are from the closed structure (though the enzyme has also been crystallized with only magnesium bound, and it adopted an open conformation).
The structure of PEPPM is very similar to that of methylisocitrate lyase, an enzyme involved in propanoate metabolism whose substrate is also a low-molecular weight carboxylic acid—the beta-barrel structure as well as the active site layout and multimerization geometry are the same. Isocitrate lyase is also quite similar, though each subunit has a second, smaller beta domain in addition to the main beta barrel.
Mechanism
Phosphoenolpyruvate mutase is thought to exhibit a dissociative mechanism.[4] A magnesium ion is involved as a cofactor. The phosphoryl/phosphate group also appears to interact ionically with Arg159 and His190, stabilizing the reactive intermediate. A phosphoenzyme intermediate is unlikely because the most feasible residues for the covalent adduct can be mutated with only partial loss of function. The reaction involves dissociation of phosphorus from oxygen 2 and then a nucleophilic attack by carbon 3 on phosphorus. Notably, the configuration is retained at phosphorus, i.e. carbon 3 of PPR adds to the same face of phosphorus from which oxygen 2 of PEP was removed; this would be unlikely for a non-enzyme-catalyzed dissociative mechanism, but since the reactive intermediate interacts strongly with the amino acids and magnesium ions of the active site, it is to be expected in the presence of enzyme catalysis.
Residues in the active-site gating loop, particularly Lys120, Asn122, and Leu124, also appear to interact with the substrate and reactive intermediate; these interactions explain why the loop moves into the closed conformation on substrate binding.
Biological function
Because phosphoenolpyruvate mutase has the unusual ability to form a new carbon-phosphorus bond, it is essential to the synthesis of phosphonates, such as phosphonolipids and the antibiotics fosfomycin and bialaphos. The formation of this bond is quite thermodynamically unfavorable; even though PEP is a very high-energy phosphate compound, the equilibrium in PEP-PPR interconversion still favors PEP.[1] The enzyme phosphonopyruvate decarboxylase presents a solution to this problem: it catalyzes the very thermodynamically favorable decarboxylation of PPR, and the resulting 2-phosphonoacetaldehyde is then converted into biologically useful phosphonates. This allows phosphoneolpyruvate's reaction to proceed in the forward direction, due to Le Chatelier's principle. The decarboxylation removes product quickly, and thus the reaction moves forward even though there would be much more reactant than product if the system were allowed to reach equilibrium by itself.
β-Galactosidase, is a glycoside hydrolase enzyme that catalyzes hydrolysis of terminal non-reducing β-D-galactose residues in β-D-galactosides.
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Malate dehydrogenase (EC 1.1.1.37) (MDH) is an enzyme that reversibly catalyzes the oxidation of malate to oxaloacetate using the reduction of NAD+ to NADH. This reaction is part of many metabolic pathways, including the citric acid cycle. Other malate dehydrogenases, which have other EC numbers and catalyze other reactions oxidizing malate, have qualified names like malate dehydrogenase (NADP+).
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Phosphopyruvate hydratase, usually known as enolase, is a metalloenzyme (EC 4.2.1.11) that catalyses the conversion of 2-phosphoglycerate (2-PG) to phosphoenolpyruvate (PEP), the ninth and penultimate step of glycolysis. The chemical reaction is:
Phosphoenolpyruvate carboxylase (also known as PEP carboxylase, PEPCase, or PEPC; EC 4.1.1.31, PDB ID: 3ZGE) is an enzyme in the family of carboxy-lyases found in plants and some bacteria that catalyzes the addition of bicarbonate (HCO3−) to phosphoenolpyruvate (PEP) to form the four-carbon compound oxaloacetate and inorganic phosphate:
Phosphoenolpyruvate carboxykinase is an enzyme in the lyase family used in the metabolic pathway of gluconeogenesis. It converts oxaloacetate into phosphoenolpyruvate and carbon dioxide.
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Bisphosphoglycerate mutase is an enzyme expressed in erythrocytes and placental cells. It is responsible for the catalytic synthesis of 2,3-Bisphosphoglycerate (2,3-BPG) from 1,3-bisphosphoglycerate. BPGM also has a mutase and a phosphatase function, but these are much less active, in contrast to its glycolytic cousin, phosphoglycerate mutase (PGM), which favors these two functions, but can also catalyze the synthesis of 2,3-BPG to a lesser extent.
Glutaryl-CoA dehydrogenase (GCDH) is an enzyme encoded by the GCDH gene on chromosome 19. The protein belongs to the acyl-CoA dehydrogenase family (ACD). It catalyzes the oxidative decarboxylation of glutaryl-CoA to crotonyl-CoA and carbon dioxide in the degradative pathway of L-lysine, L-hydroxylysine, and L-tryptophan metabolism. It uses electron transfer flavoprotein as its electron acceptor. The enzyme exists in the mitochondrial matrix as a homotetramer of 45-kD subunits. Mutations in this gene result in the metabolic disorder glutaric aciduria type 1, which is also known as glutaric acidemia type I. Alternative splicing of this gene results in multiple transcript variants.
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Riboflavin synthase is an enzyme that catalyzes the final reaction of riboflavin biosynthesis. It catalyzes the transfer of a four-carbon unit from one molecule of 6,7-dimethyl-8-ribityllumazine onto another, resulting in the synthesis of riboflavin and 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione:
In enzymology, D-lysine 5,6-aminomutase is an enzyme that catalyzes the chemical reaction
In enzymology, a phosphonopyruvate hydrolase (EC 3.11.1.3) is an enzyme that catalyzes the chemical reaction
The enzyme methylisocitrate lyase catalyzes the chemical reaction
In enzymology, a nucleoside-phosphate kinase is an enzyme that catalyzes the chemical reaction
Phosphoribulokinase (PRK) (EC 2.7.1.19) is an essential photosynthetic enzyme that catalyzes the ATP-dependent phosphorylation of ribulose 5-phosphate (RuP) into ribulose 1,5-bisphosphate (RuBP), both intermediates in the Calvin Cycle. Its main function is to regenerate RuBP, which is the initial substrate and CO2-acceptor molecule of the Calvin Cycle. PRK belongs to the family of transferase enzymes, specifically those transferring phosphorus-containing groups (phosphotransferases) to an alcohol group acceptor. Along with ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCo), phosphoribulokinase is unique to the Calvin Cycle. Therefore, PRK activity often determines the metabolic rate in organisms for which carbon fixation is key to survival. Much initial work on PRK was done with spinach leaf extracts in the 1950s; subsequent studies of PRK in other photosynthetic prokaryotic and eukaryotic organisms have followed. The possibility that PRK might exist was first recognized by Weissbach et al. in 1954; for example, the group noted that carbon dioxide fixation in crude spinach extracts was enhanced by the addition of ATP. The first purification of PRK was conducted by Hurwitz and colleagues in 1956.
ATP + Mg2+ - D-ribulose 5-phosphate ADP + D-ribulose 1,5-bisphosphate
Pyruvate, phosphate dikinase, or PPDK is an enzyme in the family of transferases that catalyzes the chemical reaction
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References
1 2 Bowman E, McQueney M, Barry RJ, Dunaway-Mariano D (1988). "Catalysis and thermodynamics of the phosphoenolpyruvate phosphonopyruvate rearrangement - entry into the phosphonate class of naturally-occurring organo-phosphorus compounds". J. Am. Chem. Soc. 110 (16): 5575–5576. doi:10.1021/ja00224a054.
1 2 Liu S, Lu Z, Jia Y, Dunaway-Mariano D, Herzberg O (2002). "Dissociative phosphoryl transfer in PEP mutase catalysis: structure of the enzyme/sulfopyruvate complex and kinetic properties of mutants". Biochemistry. 41 (32): 10270–10276. doi:10.1021/bi026024v. PMID12162742.
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