Active-site conformational motions

Molecular Dynamics (MD) Simulation of the Active-Site Conformational Motions in Forming an Active Enzyme—Substrate Complex of the Enzymatic Reaction... 

Loop motion Active site conformation adap- Nanoseconds (ns) to micro-... 

The highly conserved flexible peptide loops that close over the active sites of TIM when substrate binds - were held rigid in open conformations in the studies described. However, the dynamics of these loops may be of kinetic as well as mechanistic importance and may affect the rate constant of the reaction in two ways. First, each loop may serve as a gate to the active site, which can prevent substrate access when it is closed and may therefore reduce the rate constant. Second, each loop may serve as a scoop to guide the substrate into the active site and, thereby, increase the rate constant. To investigate whether the loops function as gates to the active sites, their motion has been simulated in the absence of substrate these simulations are outlined and discussed here. Further simulations that incorporate the motion of both the loops and the substrate are necessary to investigate whether the loops can facilitate access of the substrate to the active sites. 

The idea of conformational coupling of ATP synthesis and electron transport is especially attractive when we recall that ATP is used in muscle to carry out mechanical work. Here we have the hydrolysis of ATP coupled to motion in the protein components of the muscle. It seems reasonable that ATP should be formed as a result of motion induced in the protein components of the ATPase. Support for this analogy has come from close structural similarities of the F, ATPase P subunits and of the active site of ATP cleavage in the muscle protein myosin (Chapter 19). 

A key structural and mechanistic feature of lactate and malate dehydrogenases is the active site loop, residues 98-110 of the lactate enzyme, which was seen in the crystal structure to close over the reagents in the ternary complex.49,50 The loop has two functions it carries Arg-109, which helps to stabilize the transition state during hydride transfer and contacts around 101-103 are the main determinants of specificity. Tryptophan residues were placed in various parts of lactate dehydrogenase to monitor conformational changes during catalysis.54,59,60 Loop closure is the slowest of the motions. 

The internal motion of T4 lysozyme in the crystal was interpreted as an inter-domain motion corresponding to opening and closing of the active site cleft (Weaver et al., 1989). Hinge-bending and substrate-induced conformational transition in T4 lysozyme in solution were confirmed in a study by site-directed labelling (Mchaourban et al., 1997). 

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