Enzymatic Catalysis of Proton Transfer

Studies on proton transfer to and from carbon in model reactions have shown that the activation barrier to most enzyme-catalyzed reactions is composed mainly of the thermodynamic barrier to proton transfer (Fig. 1.1), so that in most cases this barrier for proton transfer at the enzyme active site will need to be reduced in order to observe efficient catalysis. A smaller part of the activation barrier to deprotonation of a-carbonyl carbon is due to the intrinsic difficulty of this reaction to form a resonance stabilized enolate. There is evidence that part of the intrinsic barrier to proton transfer at a-carbonyl carbon reflects the intrinsic instability of negative charge at the transition state of mixed sp -sp -hybridization at carbon [79]. Small buffer and metal ion catalysts do not cause a large reduction in this intrinsic reaction barrier. 

There is extensive evidence from site-directed mutagenesis and other studies of enzymes that catalyze proton transfer that acidic and basic amino side chains and, in some cases, metal cations, are required for the observation of efficient catalysis. However, catalysis of the deprotonation of a-carbonyl by small molecule analogs of these side chains, and by metal cations is generally weak. Relatively little attention has been directed towards understanding the mechanism for the enhancement of Bronsted acid/base and electrophilic catalysis for enzyme-catalyzed reactions 

There may also be a reduction in the intrinsic barrier for proton transfer at the enzyme active site compared to solution [80]. This possibility is intriguing however, we are unable to offer a convincing mechanism for such a reduction of intrinsic reaction barrier. 

We acknowledge the National Institutes of Health Grant GM 39754 for its generous support of the work from our laboratory described in this review. 

Vinogradov, R. H. Linnell Hydrogen Bonding, Van Nostrand-Reinhold, New York, 1971, pp. 120-124. 

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Enzymatic Catalysis of Proton Transfer at Carbon Atoms... 

R464 J. P. Richard, Enzymatic Catalysis of Proton Transfer and Decarboxylation Reactions , Pure Appl. Chem., 2011, 83, 1555. 

Histidine is the least basic of the three basic amino acids. The imidazole ring in the side chain of the free amino acid loses its proton at about pH 6. When histidine is incorporated into proteins, the pKa is raised to about 7 (Table 5.3). Becauase the histidine side chain can exchange protons near physiological pH, it often plays a role in enzymatic catalysis involving proton transfer. 

Acid-base catalysis appears to be an important factor in virtually all enzymatic reactions. The rates of proton transfer reactions have been well studied in model systems,30 but not during the course of enzyme catalysis. The protonation and deprotonation of acids and bases can be represented as... 

Despite all the problems inherent to QM/CM approaches, some extremely interesting and perceptive work has been described in the literature recently in which all sorts of approaches have been used, improvements introduced and results obtained ([351, 372] and references therein). The study of enzyme catalysed reaction mechanisms, the calculation of relative binding free energies of substrates and inhibitor, and the determination of proton transfer processes in enzymatic reactions, are all good examples of enzyme-ligand interactions studies. Even though Warshel s EVB method [349] probably remains the most practical QM/CM approach for the study of enzyme catalysis, very useful work has been reported on enzyme catalysed reactions ([381] for an excellent review-[238, 319, 382-384]). This is a consequence of the accuracy of QM to treat the active site and inhibitor/substrate and the viability of classical mechanics to model the bulk of the enzyme not directly involved in the chemical reaction. 

The first indication of the importance of proton-transfer reactions in enzyme catalysis come from the observation that the rate of most enzyme-catalysed reactions displays a relatively simply, sigmoidal or bell-shaped pH dependence. Thus enzymatic reactions require a small number of acids in a definite state of ionization. Later mechanistic studies indeed confirmed that in many cases these acids and bases - usually identifiable from the pK values of the pH-rate profile -act as proton donors and proton acceptors in the rate-limiting step of the catalytic process. Since in biological systems enzymatic reactions occur almost invariably near neutrality, where oxonium and hydroxide ion concentrations are at a minimum, it is not surprizing to find that enzymes make extensive use of general acid and general base catalysis. 

Edsall, J. T. George Scatchard, John G. Kirkwood, and the electrical interactions of amino acids and proteins. Trends Biochem. Sci. 7 (1982) 414-416. Eigen, M. Proton transfer, acid-base catalysis, and enzymatic hydrolysis. Angew. Chem. Int. Ed. Engl. 3 (1964) 1-19. 

Bronsted acid/base catalysis is the most common enzymatic mechanism, since nearly all enzymatic reactions involve a proton transfer. This means that nearly all enzymes have acidic and/or basic groups in their active site. In add catalysis, the substrate is protonated by one of the amino add residues at the active site (typically aspartic acid, glutamic acid, histidine, cysteine, lysine, or tyrosine). This residue itself must therefore be protonated at the readion pH (typically between pH 5 and 9), with a pKa just above this value. Conversely, in base catalysis, the pJCa of the deprotonating residue must be just below the physiological pH. Some enzymes can even carry out bifunctional catalysis, by protonating and deprotonating two different sites on the same substrate molecule simultaneously. 

In the course of developing the idea of the enzymatic catalysis mechanism Poltorak [99] stated the uniformity of enzymatic catalysis mechanisms in the framework of suggested notion of linear chain of bond redistribution (linear CBR). Actually, this idea laid the foundation for the catalase reaction mechanism suggested by Poltorak. In this mechanism, owing to composition of linear CBRs he showed the means for effective proton transfer between... 

In the framework of general BRC theory in the example of PPFe3+0H/Al203, the unified picture of two-proton transfer to acidic-basic groups of the carrier (A1203) with electron transfer to the active site (PPFe3+OH) is observed. Finally, the substrate is redox converted. It is typical that in enzymatic catalysis conditions without acidic-basic groups redox processes are suppressed. 

The intrinsic catalytic properties of enzymes are modified either during immobilization or after they were immobilized [25-27], In heterogeneous catalysis such as is carried out by immobilized enzymes, the rate of reaction is determined not simply by pH, temperature and substrate solution, but by the rates of proton, heat and substrate transport, through the support matrix to the immobilized enzyme. In order to estimate this last phenomenon, we have studied the internal mass transfer limitation both in hexane and in SC C02, with different enzymatic support sizes. 

The most common type of biocatalytic reactions is proton transfer (115). Nearly, every enzymatic reaction involves one or more proton-coupled steps. Transition-state proton bridging and intramolecular proton transfer (general acid-base catalysis) are important strategies to accelerate substrate conversion processes. Moreover, proton transfer also plays a fundamental role in bioenergetics (116). 

As discussed, AdoMet is a high energy compound, and therefore, AdoMet-dependent methylation reactions are known to be irreversible. However, enzymatic catalysis is often required to enhance the rate of the reaction. Enzymes employ a variety of ways to enhance the nucleophilicity on the attacking atom in a substrate. Often, the reaction results in a proton exchange for the methyl group. The proton can be removed before, in concert with, or after the methyl transfer this step usually requires the presence of a general base in the active site. 

Eigen has also been interested in the chemical basis of the origin of life, which seems to be a very different question from his previous studies, in reality it is not. Two of his papers were named Citation Classics by Current Contents. One paper was Proton-transfer, acid-base catalysis, and enzymatic hydrolysis. Part I elementary processes. An ew. Chem. 1963, 75, 489 Int. Ed. En l. 1964, 5, 1-19. It was one of the most cited papers in the field. The German and the English versions have been cited more than 285 and 965 times, respectively, by 1990. Then Eigen worked on evolution and published another paper, Self-organization of matter and the evolution of biological macromolecules. Naturwissenschafien 1971, 58, 465-523. This paper was the most cited paper in that journal in over 490 publications by 1990. 

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