Glutamine synthetase reaction mechanism

The above data, along with other data from Meister s group (102, 103), provided the basis for computer modeling studies (104) of the active site conformations of substrates and intermediates in the glutamine synthetase reaction. The conclusions are that both structures II and III are in the reaction mechanism. 

Meister has more recently studied the glutamine synthetase reaction in considerable detail, and has proposed the following mechanism ... 

The kinetic reaction mechanism appears to be random, and for the reaction to proceed, all substrates must reside as a E-D-Ala-D-Ala-MgATP quaternary complex. Except for its activation of an a-carboxylate to form a peptide bond, the enzyme s mechanism appears to be completely analogous to that catalyzed by glutamine synthetase, which forms a y-glutamyl-phosphate intermediate. There is strong evidence for the participation... 

A three-substrate, three-product enzyme-catalyzed reaction scheme in which the three substrates (A, B, and C) and three products (P, Q, and R) can bind to and be released in any order. A number of enzymes have been reported to have this mechanism for example, adenylosuccinate synthetase , glutamate dehydrogenase, glutamine synthetase , formyltetrahydrofolate synthetase, and tubulin tyrosine ligase . See Multisubstrate Mechanisms... 

Since the reaction catalyzed by glutamine synthetase has three substrates (L-glutamate, NH3, and metal-ATP) and three products (L-glutamine, P , and metal-ADP), the kinetic mechanism as deduced by steady-state kinetics... 

This section summarizes recent data obtained from biophysical techniques on the nature of the metal ion, catalytic, and modifier sites of E. coli glutamine synthetase. Distance relationships between these sites are given in Fig. 27. This enzyme has many diverse modes of biochemical control. Recent data presented in this review demonstrate that progress is being made toward understanding the structural basis for the control of enzymic catalysis. When x-ray data become available 127), the amino acid groups at the catalytic site will be known and will provide further details concerning the mechanism of the enzymic reaction. 

S H Liaw, D Eisenberg (1994) Structural model for the reaction mechanism of glutamine synthetase, based on five crystal structures of enzyme-substtate complexes, Biochemistry 33(3) 675—681... 

Rhee, S. G., Chock, P. B., and Stadtman, E. R. (1976). Mechanistic studies of glutamine synthetase from Eschericihia coli. An integrated mechanism for biosynthesis, transferase, ATPase reactions. 

Fig. 10. Active site of glutamine synthetase. The two metal ions are designated ni and nj and represent their location in the crystal stmcture (98). The metal ions coordinate the substrates glutamate (ni) and ATP (n ) and are considered to be involved in the mechanism as shown. The ni site serves to orient the glutamate for attack on ATP and the 02 metal ion aids in making ADP a good leaving group. The reaction shown represents the initial step in the overall mechanism. Subsequent steps involve attack of NH3 on the y-glutamyl-P intermediate.

The mechanism of action of glutamine synthetase has been extensively studied, and both the bacterial and mammalian enzymes were reviewed in Volume X of this series (83, 84). Many experimental approaches have been applied to the question of the chemical activation of the y-carboxyl group of glutamate and its reaction with ammonia. All of these experiments supported the stepwise chemical mechanism outlined in reactions (24a) and (24b) (8J),... 

To examine the kinetic competence of y-glutamyl phosphate formation at the active site of glutamine synthetase, the technique of positional isotope exchange (PIX) was introduced to studies of the mechanism of phospho transfer (85). This experiment circumvented the problem of the instability of the intermediate, which complicated direct kinetic experiments. In the PIX experiment, the rate at which glutamine synthetase catalyzes reaction (25) was measured. 

Describe the cumulative feedback control of glutamine synthetase from E. coli. Explain the mechanisms and functions of the reversible covalent modifications and describe the advantage of employing an enzymatic cascade in regulating this reaction. 

Wedler and Boyer (1972) were however unable to obtain any evidence for a sequential reaction mechanism for E. coli glutamine synthetase. Under their experimental conditions the enzyme failed to carry out any partial reaction unless all the substrates were present. They concluded that the reaction proceeds by a concerted mechanism with random substrate binding. Later work (Wedler and Horn, 1976) suggests that the mammalian enzyme... 

Glutamine is synthesized in pigeon liver homogenates from glutamic acid and ammonia. The reaction requires ATP and a specific enzyme, glutamine synthetase. ADP is formed and inorganic phosphate is released in the reaction. The mechanism of the reaction is similar to that involved in glutathione biosynthesis. 

Glutamine synthetase was found in the microsomes and the supernatant of rat brain and liver. The microsomal enzyme is weakly bound to the endoplasmic reticulum. The sheep enzyme has been purified and has a molecular weight of 450,000. Glutamine is bound to the enzyme in at least four bindings sites. The postulated mechanism of the enzymic reaction involves (1)... 

However, almost none of appropriate enzyme models have ever been reported for the reaction path control or the allosteric control in homogeneous aqueous solution(2). In biological systems, both control mechanisms are very frequently and significantly operating as in the case of the reaction path control of the amino acid metabolism by individual enzyme(3) or the allosteric control of levels of many important compounds by glutamine synthetase. 

Glutamine synthetase, the enzyme catalysing this reaction, has been purified from sheep brain . The possible mechanism of its action has been reviewed by Meister, but none of the intermediate steps. 

Glutamine (and probably asparagine) can be destroyed by two independent routes. The enzyme catalysing direct deamidation to glutamate is activated by phosphate, suggesting a mechanism related to that of glutamine synthetase. The second path involves transamination (equation 30) and subsequent deamidation (equation 31) of a-ketoglutaramic acid. The two reactions are catalysed by separate... 

Other naturally occurring compounds that have been shown to inhibit plant glutamine synthetase are oxetin (Omura et al., 1984) and 5-hydroxylysine (Leason et al., 1982 Acaster and Weitzman, 1985). Two additional molecules, 2-amino-4-phosphonobutyric acid and 3-amino-(3-carboxypropyl)-(phos-phonomethyl)phosphinic acid, have been specifically synthesized to study the reaction mechanisms of glutamine synthetase (Lea and Ridley, 1989). There are numerous reports in the literature that MSO or PPT can cause a massive build-up of ammonia in leaves in the light (see Lea and Ridley, 1989, for a full list) and severely disrupt amino acid metabolism (Rhodes et al., 1986). It has... 

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