Enzyme transition states from theory and experiment.

Understanding how enzymes catalyze specific reactions has yielded important insights into biology, provided the necessary context for the design of novel catalysts, and has facilitated the development of enzyme inhibitors for therapeutic purposes. The contributions of Professor W. Wallace Cleland helped to forge the modern framework for steady state enzyme kinetics, and together with his colleagues he made pioneering contributions to establishing heavy atom isotope effect analyses as a powerful approach for investigating the chemical mechanisms of enzymecatalyzed reactions. Professor Cleland collaborated with and trained numerous talented enzymologists and generations of scientists. Several facets of W.W. Cleland's contributions to science are illustrated and reflected in this special issue that focuses on using kinetic isotope effects to understand chemical and enzymatic reaction mechanisms. Along with experimental advances, computation has grown and developed as an essential partner in the interpretation of kinetic isotope effects (KIEs). Transition state theory poses that kinetic isotope effects reveal bonding changes in the transition state and thus the interpretation of KIEs in terms of transition state structure is complicated by kinetic ambiguity KIEs reflect changes in all vibrational modes between the ground state and transition state that involve the atom being analyzed. Therefore, measured KIEs typically reflect contributions frommore than one of thesemodes, or fromnewmodes introduced by interactionswith a catalyst. As illustrated in the contributions to this issue, the combination of computational and experiment provides the ability to estimate themagnitude of different contributions to observed KIEs and therefore to evaluate alternative mechanisms and to derive transition state structures corresponding to competing pathways. One of W. W. Cleland's significant contributions is in the development of steady state enzyme kinetics. His description of internal competition kinetics used to calculate kinetic isotope effects and commitment factors is widely used. Building on this framework, Dr. Vernon Anderson provides a review and derivation of the equations for alternative substrate kinetics. This contribution further includes a detailed description of the expansion of internal competition kinetics to analyze the relative rate constants of large numbers of substrates, which has great promise in high throughput quantitative biochemistry. Importantly, the quantitative framework includes determination of equations for calculation of relative rate constant using both precursor and product ratios, and therefore is a resource adaptable to many enzyme systems.