Phosphoryl transfer enzymes

Simple additions of such labile aquo ions as Mg, Ca, Zn and Mn "1", which are of importance in enzymic phosphoryl transfer, have resulted in only very modest catalytic effects for reactions of phosphate species (3). The much more effective t Com unit possesses the special advantage that it remains intact for long periods, while trans/cis isomerization and substitution in the fifth and sixth coordination sites proceed at moderately rapid and generally convenient rates. These characteristics make it particularly suitable for use in model studies (4). 

Although the possibility of pseudorotation during enzymic phosphoryl transfer has been raised many times, there is as yet no unambiguous example of it. If groups enter and leave from apical positions, the trigonal bipyramidal intermediate may have a lifetime less than the period of a vibration and thus be too unstable to exist. In practice, therefore, the rule retention = double displacement, inversion = single displacement appears to hold for phosphoryl transfer at least as well as for glycosyl transfer. 

A series of kinetic experiments reported by Herschlag and Jencks (7) addressed the influence of metal ions on phosphoryl transfer reactions for non-enzymic phosphoryl transfer between pyridine and carboxylate ions (and other nucleophiles). The reaction of actetate ion with phosphorylated -y-picoline monoanion and the reverse reaction of y-picoline with acetyl phosphate dianion are... 

A recent kinetic study of the catalysis of hydrolysis of phosphorylated pyri-dines by Mg in aqueous solution as a model for enzymic phosphoryl transfer reactions has been reported by Herschlag and Jencks (23). They have provided evidence that a l(y -10 rate enhancement can be obtained in the presence of Mg +. From the analysis, approximately a l(y -fold rate enhancement can be attributed to the greater nucleophilicity of the species Mg(OH)+ compared to... 

The structure and function of phosphoproteins has provoked much interest in the past year. The application of n.m.r. to the study of protein structure and function has been reviewed while the stereochemistry of enzymic phosphoryl transfer and the enzymology of kinases have been described in recent volumes of Methods in Enzymology. The identification of the sites of phosphorylation on phosphoproteins is an important prerequisite in attempting the elucidation of the functions of these compounds and methods have been described for the detection of 0-phosphoserine and 0-phosphotyrosine residues in proteins. The enzymic phosphorylation and dephosphorylation of tyrosine residues in proteins has been investigated, and a simple synthesis of O-phospho-L-tyrosine from the amino acid and a phosphoric oxide/orthophosphoric acid mixture has been described. ... 

The detection and characterization of intermediates in complex reactions is often accomplished through the use of isotope-exchange studies. In the case of enzymic phosphoryl transfer reactions, the presence of a phosphorylated enzyme intermediate is implicated by a partial exchange process as exemplified by the following reaction of hexokinase (Knowles, 1980) ... 

Metal ions are generally required for enzymic phosphoryl transfer. Their catalytic effects on phosphate ester and amide hydrolysis in model systems are, however, quite modest in the absence of an Intramolecular binding site. Given the latter, large accelerations are seen. An o-imldazolyl function binds Cu" as in to yield a greater than 10 x... 

Herschlag, D., Jencks, W.P. Catalysis of the hydrolysis of phosphorylated pyridines by Mg(OH)L a possible model for enzymic phosphoryl transfer. Biochemistry 1990, 29(21), 5172-5179. 

FIGURE 10.26 Glucose transport in E. coli is mediated by the PEP-dependent phosphotransferase system. Enzyme I is phosphorylated in the first step by PEP. Successive phosphoryl transfers to HPr and Enzyme III in Steps 2 and 3 are followed by transport and phosphorylation of glucose. Enzyme II is the sugar transport channel. 

FIGURE 10.27 The path of the phosphoryl group through the PTS mechanism. Reactive phosphohistidine intermediates of Enzyme I, HPr, and Enzyme III transfer phosphoryl groups from PEP to the transported sugar. 

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... 

Synthetic nonhydrolyzable analogs of nucleoside triphosphates (Figure 33-13) allow investigators to distinguish the effects of nucleotides due to phosphoryl transfer from effects mediated by occupancy of allosteric nucleotide-binding sites on regulated enzymes. 

This can be used in several ways. H+-ATPase plays a key role in oxidative phosphorylation (ATP synthesis) as it transfers protons back into the matrix space with simultaneous synthesis of ATP (with temporary enzyme phosphorylation, cf. page 451). 

The AMl/d-PhoT model [33] is a parameterization of a modified AMl/d Hamiltonian developed specifically to model phosphoryl transfer reactions catalyzed by enzymes and ribozymes for use in linear-scaling calculations and combined QM/MM simulations. The model is currently parametrized for H, O, and P atoms to reproduce... 

Mechanistic studies on enzyme-catalyzed phosphoryl transfer, 40, 49... 

Magnesium has its role intimately intertwined with phosphate in many phosphoryl transfer reactions, as Mg-ATP in muscle contraction, in the stabilization of nucleic acid structures as well as in the catalytic activity of ribozymes (catalytic RNA molecules). It also serves as a structural component of enzymes, and is found as the metal centre in chlorophylls, which absorbs light energy in photosynthesis. 

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. 

Knowles, J.R. (1980) Enzyme-catalysed phosphoryl transfer reactions, Annu. Rev. Biochem., 49, 877-919. 

F. Hollfelder, D. Herschlag, The Nature of the Transition State for Enzyme-Catalyzed Phosphoryl Transfer. Hydrolysis of O-Aryl Phosphorothioates by Alkaline Phosphatase , Biochemistry 1995, 34, 12255-12264. 

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. 

This enzyme [EC 2.7.1.32] catalyzes the phosphoryl transfer from ATP with choline to produce ADP and O-phosphocholtne. Ethanolamine and its methyl and ethyl derivatives can also serve as substrates. 

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