Substrate binding concerted mechanism

There is still a third possible mechanism for the fumarate hydratase reaction. The proton and hydroxyl groups may be added simultaneously in a concerted reaction. However, observed kinetic isotope effects are not consistent with this mechanism. In 1997 the structure of fumarase C of E. coli was reported. Each active site of the tetrameric enzyme is formed using side chains from three different subunits. The H188 imidazole is hydrogen bonded to an active site water molecule and is backed up by the E331 carboxy-late which forms a familiar catalytic pair. However, these results have not clarified the exact mode of substrate binding nor the details of the catalytic mechanism. Structural studies of fumarate hydratase from yeast and the pig are also in progress. 

In addition to the cationic, radical, and non-synchronous concerted mechanisms outlined above, many other proposals have been offered [9, 56], In a recent provocative paper, an organometallic mechanism was postulated for the activation of alkanes by a ferryl porphyrin model species [79]. Less reactive substrates such as H2, D2 and CH4 were observed to inhibit the reaction between the synthetic catalyst and cyclohexane. In the proposed mechanism, a 2 -t- 2 C-H addition across the Fe-O bond is preceded by coordination of the alkane to the metal center to form an intermolecular <7-adduct. Inhibition arises from preferential binding of the smaller substrates to the congested metal site. Attempts to identify a similar effect with sMMO have been unsuccessful the presence of H2 had no effect on the rate of reaction between methane and Q (A. M. Valentine, S. S. Stahl, S. J. Lippard, unpublished results). 

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... 

The basic kinetic properties of this allosteric enzyme are clearly explained by combining Monod s theory and these structural results. The tetrameric enzyme exists in equilibrium between a catalytically active R state and an inactive T state. There is a difference in the tertiary structure of the subunits in these two states, which is closely linked to a difference in the quaternary structure of the molecule. The substrate F6P binds preferentially to the R state, thereby shifting the equilibrium to that state. Since the mechanism is concerted, binding of one F6P to the first subunit provides an additional three subunits in the R state, hence the cooperativity of F6P binding and catalysis. ATP binds to both states, so there is no shift in the equilibrium and hence there is no cooperativity of ATP binding. The inhibitor PEP preferentially binds to the effector binding site of molecules in the T state and as a result the equilibrium is shifted to the inactive state. By contrast the activator ADP preferentially binds to the effector site of molecules in the R state and as a result shifts the equilibrium to the R state with its four available, catalytically competent, active sites per molecule. 

The methanolyses of several phosphate/phosphonate esters and their thio analogues [e.g. 0,0-diethyl 0-(4-nitrophenyl) phosphate, paraoxon (101 X = O, Z = 4-N02), 0,0-diethyl S- S-dichlorophenyl) phosphorothioate, (102 R = OEt), and 0-ethyl S-OA-dichlorophenyl) methylphosphonothioate (102 R = Me)] catalysed by methoxide and the complex of Zn2+ (MeO-) with 1,5,9-triazacyclododecane (79 M = Zn) were studied in methanol at 25 °C. The reaction of methoxide and (79 M = Zn) with the entire series of esters appears to adhere to a common mechanism that involves pre-equilibrium binding of the substrate, followed by intramolecular attack of the coordinated methoxide concerted with OAr or SAr leaving group departure.79... 

Fig. is. Model for the mechanism of homotropic cooperativity in aspartate transcarbamylase. Shown schematically are the two extreme conformations in the T state and the R state. The binding of the substrates at one active site induces the domain closure in that catalytic chain and requires a quaternary conformational change which allows the 240s loops of the upper and lower catalytic chains to move to their final positions. The formation of the R state, in a concerted fashion, is further stabilized by a variety of new interactions as shown. [Reprinted with permission from Ref. (/).]... 

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