Phosphotransferase donors

Several unique features distinguish the phosphotransferase. First, phos-phoenolpyruvate is both the phosphoryl donor and the energy source for sugar transport. Second, four different proteins are required for this transport. Two of these proteins (Enzyme I and HPr) are general and are required for the phosphorylation of all PTS-transported sugars. The other two proteins (Enzyme II and Enzyme III) are specific for the particular sugar to be transported. 

An important additional pathway is indicated in reactions I and II of Figure 47-9. This involves enzymes destined for lysosomes. Such enzymes are targeted to the lysosomes by a specific chemical marker. In reaction I, a residue of GIcNAc-1-P is added to carbon 6 of one or more specific Man residues of these enzymes. The reaction is catalyzed by a GIcNAc phosphotransferase, which uses UDP-GlcNAc as the donor and generates UMP as the other product ... 

This phosphotransferase [EC 2.7.1.1] catalyzes phosphorylation of D-glucose as well as many other aldo- and keto-hexoses (with the notable exception of o-galac-tose) MgATP is the active phosphoryl donor, and the overall reaction is highly favorable T eq = [ADP]... 

Attachment of phosphopantetheine to proteins is catalyzed by a phosphotransferase that utilizes CoA as the donor. A phosphodiesterase removes the phosphopantetheine, providing a turnover cycle.15, 5b A variety of synthetic analogs have been made.4 16 The reactive center of CoA and phosphopantetheine is the SH group, which is carried on a flexible arm that consists in part of the (3-alanine portion of pantothenic acid. A mystery is why pantoic acid, a small odd-shaped molecule that the human body cannot make, is so essential for life. The hydroxyl group is a potential reactive site and the two methyl groups may enter into formation of a "trialkyl lock" (p. 485), part of a sophisticated "elbow" or shoulder for the SH-bearing arm. 

As discussed briefly in Section I,A, glucose-6-phosphatase is now known to be a multifunctional enzyme capable of catalyzing potent phosphotransferase as well as phosphohydrolase reactions [see Eqs. (1)—(4) ]. Compounds demonstrated to function as effective phosphoryl donors include fructose-6-P (30), mannose-6-P (40), PPi (35-38), a variety of nucleosidetriphosphates and nucleosidediphosphates—most effectively CTP, CDP, deoxy-CTP, ATP, ADP, GTP, GDP, and ITP (41, 45)— carbamyl-P (43), phosphoramidate (44), phosphopyruvate (42, 43) and glucose-6-P itself (30, 31). The various phosphoryl donors are also hydrolyzed by action of the enzyme (see preceding references). Eqqa-tions (1)—(4), which describe these various activities, are given in Section I,A. 

A Km value of 2.8 mM for phosphoramidate serving as a phosphoryl donor with untreated rat liver microsomes has been observed at pH 6.5 by Parvin and Smith (44), who also reported values of approximately 0.45 for the ratio of phosphoramidate-glucose phosphotransferase/PPi-glucose phosphotransferase with both rat liver and kidney microsomes. The latter assays were carried out at pH 6.5 with 0.4 M glucose and 20 mM phosphate substrates. 

Fig. 6. Effects of varied substrate concentrations on velocity v of phosphotransferase reactions, described in terms of conventional (136) double-reciprocal plots. In (A) v has been assessed as a function of concentration of phosphoryl donor D with phosphoryl acceptor constant at concentrations Ai, A2, Aj, or A,. In (B) v has been determined as a function of varied concentrations of phosphoryl acceptor A with phosphoryl donor held constant at concentrations Di, D2, Da, or D.. (Generalized from references Jfi, 41, and 43-)...

Nucleotides (10, 103) Various nucleoside di- and triphosphate compounds inhibit glucose-6-P phos-phohydrolase activity competitively over a broad range of pH (pH 5-8). Also function as phosphoryl donors in phosphotransferase reactions. Effects are markedly potentiated by various detergents... 

In many prokaryotes, PolyP is a direct phosphorus donor for biochemical reactions due to the action of enzymes such as polyphosphate-glucose phosphotransferase and NAD kinase. Polyphosphate kinases and PolyP AMP phosphotransferase link nucleoside-polyphosphate and inorganic PolyP. Polyphosphate kinases 1 and 2 can use PolyPs for the synthesis of different nucleoside triphosphates. 

Purine nucleotides are probably best made by phosphorylation of the corresponding nucleosides. In recent years phosphoryl chloride has proved most convenient for this purpose and may be used with an unprotected nucleoside to give yields of about 50% (B-78MI40903, p. 827). Yields of purine nucleotides up to 60% may also be obtained using a phosphotransferase and a suitable phosphate donor with the unprotected nucleoside. Typical preparations of this type using an enzyme from wheat shoots and p-nitrophenyl phosphate as a phosphate donor have been described (B-78MI40903, p. 955). 

An active phosphotransferase, requiring ATP and Mg, and catalyzing the phosphorylation of -nitrophenol has been demonstrated in the lOO.OOOg supernatants of gut tissues of the Madagascar cockroach (Gromphadorhina portentosa), tobacco hornworm (M. sexta) larvae and in whole houseflies (H. domestica) (21,30) in K. portentosa it appears to constitute a major pathway for phenol metabolism (21,30). Although the mechanism of phosphorylation has not been elaborated, it is probable that ATP serves as the direct donor of the phosphate moiety transferred to the phenol (Figure 5). 

A Phosphomutases Phosphoryl group transfer, for which the acceptor is another functional group on the donor molecule. 2.7.5. 5.4.2. Phosphomutases Intramolecular phosphotransferases... 

Phosphotransfera- ses the acceptors. Compounds different than nucleoside triphosphates are the donor and some other molecules than H20 are the acceptors. 2.7.4. Phosphotransferases with a phosphate group as acceptor... 

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