Enzyme-catalyzed reactions transition state structures

In this chapter we have seen that enzymatic catalysis is initiated by the reversible interactions of a substrate molecule with the active site of the enzyme to form a non-covalent binary complex. The chemical transformation of the substrate to the product molecule occurs within the context of the enzyme active site subsequent to initial complex formation. We saw that the enormous rate enhancements for enzyme-catalyzed reactions are the result of specific mechanisms that enzymes use to achieve large reductions in the energy of activation associated with attainment of the reaction transition state structure. Stabilization of the reaction transition state in the context of the enzymatic reaction is the key contributor to both enzymatic rate enhancement and substrate specificity. We described several chemical strategies by which enzymes achieve this transition state stabilization. We also saw in this chapter that enzyme reactions are most commonly studied by following the kinetics of these reactions under steady state conditions. We defined three kinetic constants—kai KM, and kcJKM—that can be used to define the efficiency of enzymatic catalysis, and each reports on different portions of the enzymatic reaction pathway. Perturbations... 

Secondary isotope effects measure transition-state structure 37 Quantum tunneling in enzyme-catalyzed reactions breakthroughs 42 Experimental phenomenology of quantum tunneling in enzyme-catalyzed... 

When an ionic organic reaction (the kind catalyzed by most enzymes) occurs a nucleophilic center joins with an electrophilic center. We use arrows to show the movement of pairs of electrons. Tire movement is always away from the nucleophile which can be thought of as "attacking" an electrophilic center. Notice the first step in the second example at right. The unsaturated ketone is polarized initially. However, this is not shown as a separate step. Rather, the flow of electrons from the double bond, between the a- and (1-carbons into the electron-accepting C=0 groups, is coordinated with the attack by the nucleophile. Dotted lines are often used to indicate bonds that will be formed in a reaction step, e.g., in an aldol condensation (right). Dashed or dotted lines are often used to indicate partially formed and partially broken bonds in a transition state, e.g., for the aldol condensation (with prior protonation of the aldehyde). However, do not put arrows on transition state structures. 

I n chapter 7 we saw that enzymes can increase the rates of reactions by many orders of magnitude. We noted that enzymes are highly specific in the reactions they catalyze and in the particular substrates they accept. In this chapter we explore the mechanisms of several enzyme-catalyzed reactions in greater detail. Our goal is to relate the activity of each of these enzymes to the structure of the active site, where the functional groups of amino acid side chains, the polypeptide backbone, or bound cofactors must interact with the substrates in such a way as to favor the formation of the transition state. We explore enzyme catalytic mechanisms in many subsequent chapters as well but usually in less detail than here. 

Enzymes accelerate reactions by stabilizing the transition states, the highest energy species on the reaction pathway, and thereby decreasing the activation barrier. In other words, the combination of enzyme and substrate creates a new reaction pathway whose transition-state energy is lower than it would be if the reaction were taking place without the participation of the enzyme. Enzymes have evolved to bind the transition states of substrates more strongly than the substrates themselves. Therefore, compounds that mimic the structure of the transition state are often potent inhibitors of the enzyme-catalyzed reaction. 

The activating cation presumably acts by helping the protein to maintain a productive conformation, and examples are provided by structural studies of enzymes that exhibit this property. Pauling suggested that complementarity between the stracture of the enzyme active site and the transition state of the reaction is responsible for the lowering of activation energies of enzyme-catalyzed reactions. Cation activation may aid this by ensuring that the active site has the correct... 

The enthalpy difference between these alternative reaction pathways (<2 kcal/mol) is insufficient to define a preferred transition state structure. Thus, no firm inference regarding the conformation of the intermediate in the enzyme catalyzed reaction may be drawn on the basis of the molecular orbital calculations alone. In principle, the two pathways should differ... 

Despite the amount of data and the simplicity of the chemical reaction catalyzed, the molecular basis of the catalytic mechanism of PPIases and APIases is still only poorly understood [155]. The considerable degree of amino acid sequence dissimilarity between the subgroups of peptide bond cis-trans isomerases also raises the challenging question of the mechanistic relatedness among the enzymes. At present there is a lack of detailed mechanistic investigations on APIases and multidomain PPIases. Thus, prototypic PPIases of all three families serve as the bases for unraveling catalytic pathways. One or more potential transition-state structures for enzyme-catalyzed prolyl isomerizations, alone or in combination, are consistent with the acceleration of the spontaneous rate of prolyl isomerization (Fig. 10.4). 

The information within an enzyme s active site (its shape and charge distribution) constrains the motions and allowed conformations of the substrate, making it appear more like the transition state. In other words, the information in the structure of the enzyme is used to optimally orient the substrate. As a result of this information transfer, the energy of the enzyme-substrate complex becomes closer to the AG, which means that the energy needed for the reaction to proceed to product is reduced. Consequently there is an increase in the rate of the enzyme-catalyzed reaction. Other factors, such as electrostatic effects, general acid-base catalysis, and covalent catalysis (discussed on pp. 177-180), also contribute to the increased rates of enzyme-catalyzed reactions over non-enzyme catalyzed reactions. 

Enzymes are biological catalysts. They enhance reaction rates because they provide an alternative reaction pathway that requires less energy than an uncatalyzed reaction. In contrast to some inorganic catalysts, most enzymes catalyze reactions at mild temperatures. In addition, enzymes are specific to the types of reactions they catalyze. Each type of enzyme has a unique, intricately shaped binding surface called an active site. Substrate binds to the enzyme s active site, which is a small cleft or crevice in a large protein molecule. In the lock-and-key model of enzyme action, the structures of the enzyme s active site and the substrate transition state are complementary. In the induced-fit model, the protein molecule is assumed to be flexible. 

It has been shown that both the catalyzed and the uncatalyzed reaction proceed through a chairlike transition state, stabilized in polar media59 60-143 262-263. Compound 1, an analog of the transition-state structure, proved to be a potent inhibitor of E. coli chorismate mutase-prephenate dehydrogenase204-255. For a discussion of the mechanism and structural requirements of the enzyme see refs 266 and 267. 

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