Kinase phosphoryl transfer

An example of a random, single-displacement mechanism is seen in the enzyme creatine kinase, a phosphoryl-transfer enzyme that uses ATP as a phosphoryl... 

FIGURE 19.29 A mechanism for the pyruvate kinase reaction, based on NMR and EPR studies by Albert Mildvan and colleagues. Phosphoryl transfer from phosphoenolpyrnvate (PEP) to ADP occurs in four steps (a) a water on the Mg ion coordinated to ADP is replaced by the phosphoryl group of PEP (b) Mg dissociates from the -P of ADP (c) the phosphoryl group is transferred and (d) the enolate of pyruvate is protonated. (Adapted from Mildvan, A., 1979. Advances in Eiizymology 49 103-126.)... 

Phosphoryl group transfer reactions add or remove phosphoryl groups to or from cellular metabolites and macromolecules, and play a major role in biochemistry. Phosphoryl transfer is the most common enzymatic function coded by the yeast genome and, in addition to its importance in intermediary metabolism (see Chapter 5), the reaction is catalysed by a large number of central regulatory enzymes that are often part of signalling cascades, such as protein kinases, protein phosphatases, ATPases and GTPases. 

Chen, G., Porter, M.D., Bristol, J.R., Fitzgibbon, M.J., and Pazhanisamy, S., Kinetic mechanism of the p38-alpha MAP kinase phosphoryl transfer to synthetic peptides, Biochemistry, 39, 2079, 2000. 

This phosphotransferase [EC 2.7.2.1] catalyzes the thermodynamically favored phosphorylation of ADP to form ATP Aeq = [ATP][acetate]/ [acetyl phosphate] [ADP] = 3000). GDP is also an effective phosphoryl group acceptor. This enzyme is easily cold-denatured, and one must use glycerol to maintain full catalytic activity. Initial kinetic evidence, as well as borohydride reduction experiments, suggested the formation of an enzyme-bound acyl-phosphate intermediate, but later kinetic and stereochemicaT data indicate that the kinetic mechanism is sequential and that there is direct in-line phosphoryl transfer. Incidental generation of a metaphosphate anion during catalysis may explain the formation of an enzyme-bound acyl-phosphate. Acetate kinase is ideally suited for the regeneration of ATP or GTP from ADP or GDP, respectively. 

One of the downstream elements of the signal transduction pathway, MAPKK (or, MEK) is phosphorylated by a Ser/Thr protein kinase, known as Raf, and phosphorylated MAPKK then catalyzes phosphoryl transfer to MAP kinase. The following is a recent review on the molecular and physical properties of this phosphotransferase (or, MAP kinase kinase). 

Any of a broad class of phosphoryl-transfer enzymes [EC 2.7.1.x] that catalyze the ATP-dependent phosphorylation of proteins, most often occurring at seryl, threo-nyl, and tyrosyl residues. These enzymes are central participants in cellular signal transduction pathways, and their discovery and recognition as primary control components of the cell culminated in the award of the 1992 Nobel Prize in Medicine and Physiology to American enzymologists Edwin Krebs and Edward Fischer. There is reason to believe that approximately 2% of the coding sequences in the human genome specify some 2000 different kinases that phosphorylate protein substrates. The prototypical enzyme is known as 3, 5 -cAMP-stimulated protein kinase (or, protein kinase A). See specific protein kinase... 

Both ATP and the phosphoryl acceptor become reversibly and selectively bound to the enzyme during catalysis. So far, kinases that have been shown to react by direct phosphoryl transfer between ATP and the co-substrates show strict inversion of configuration at phosphorus, while those with a phosphorylated enzyme Intermediate show retention of configuration at phosphorus (1,2). 

Schneider, B. Babolat, M. Xu, Y.W. Janin, J. Veron, M. Deville-Bonne, D. Mechanism of phosphoryl transfer by nucleoside diphosphate kinase pH dependence and role of the active site Lysl6 and Tyr56 residues. Eur. J. Biochem., 268, 1964-1971 (2001)... 

D. Herschlag, D. Nucleophilic activation by positioning in phosphoryl transfer catalyzed by nucleoside diphosphate kinase. Biochemistry, 38, 4701-4711 (1999)... 

ATP + (d)CMP = ADP + (d)CDP (<4> formation of a ternary complex, addition of substrates is random [5] <1> reaction proceeds by a sequential mechanism, a ternary complex of the enzyme with both substrates is formed as the central intermediate in the reaction [12] <3> reaction mechanism is sequential and nonequilibrium in nature, substrates bind to the enzyme in a random order, substrate binding is cooperative [14] <7> the mechanism is analogous to the phosphoryl transfer mechanism in cAMP-dependent protein kinase that phosphorylates the hydroxyl groups of serine residues [16] <8> random bi-bi mechanism [17])... 

The A isomer of /3,-y-bidentate CrATP is the only form of this complex which activates the phosphoryl-transfer reaction of pyruvate kinase.281... 

I n this way we have shown that phosphoryl transfer catalysed by Bacillus stearothermophilus and rabbit skeletal muscle phosphofructokinase (6), and rabbit skeletal muscle pyruvate kinase occurs with inversion of configuration at phosphorus (7). The simplest interpretation of these stereochemical results is that phosphoryl transfer occurs by an in-line mechanism in the enzyme substrate ternary complexes. Stereochemical analysis is thus proving to be of considerable importance for delineating the mechanism adopted by phosphokinases. ... 

Class 2. Transferases transfer chemical groups from one molecule to another, or within a single molecule. They include amino, acyl, methyl, glucosyl, and phosphoryl transferases, kinases, phosphomutases, transaldolase, and transketolase. 

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