On the Generation of Catalytic Antibodies by Transition State Analogues

Barbany, Montserrat; Gutiérrez‐de‐Terán, Hugo; Sanz, Ferrán; Villà­-Freixa, Jordi; Warshel, Arieh

doi:10.1002/cbic.200390048

Cited by 36 publications

(33 citation statements)

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“…For many natural enzyme systems, the barrier reduction (ΔΔG ‡ = ΔG 2 + ΔG 3 ) as calculated from kinetics is more dramatic than the measured differential binding to a transition-state analog (ΔG 2 ). Additionally, in the pursuit of catalytic antibodies and designed enzymes, a high affinity for transition-state analogs (ΔG 2 ) has not necessarily translated to catalytic competence, ΔΔG ‡ (20,52). We posit that some of these discrepancies could be due to ΔG 3 for systems that involve proton-transfer chemistry.…”

Section: Discussionmentioning

confidence: 98%

“…Indeed, the fact that rate ratios for enzymes can sometimes be accurately calculated by computational methods such as empirical valence bond (16,17) or quantum mechanics/molecular mechanics simulation (18), but are nearly always strikingly larger than empirically determined binding ratios, is a persistent puzzle and has been met with a number of different reactions. Many have conceded (and in some cases, shown) that inhibitors are not equivalent to the transition states of chemical reactions but rather are only crude simulacra (10,12,19,20). Others have proposed that there are dynamical contributions to catalysis that cannot be recapitulated in an equilibrium thermodynamic picture (21-23).…”

mentioning

confidence: 99%

“…2C) is that the enzymecatalyzed reaction would possess an activation barrier of 14.6 kcal·mol -1 -which is significantly higher than that observed. This disagreement is in fact very common and easy to rationalize on account of the fact that the inhibitor may only roughly recapitulate the energetics associated with the true transition state (19,20).…”

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confidence: 99%

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Thermodynamic framework for identifying free energy inventories of enzyme catalytic cycles

Fried

Boxer

2013

Proc. Natl. Acad. Sci. U.S.A.

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Pauling's suggestion that enzymes are complementary in structure to the activated complexes of the reactions they catalyze has provided the conceptual basis to explain how enzymes obtain their fantastic catalytic prowess, and has served as a guiding principle in drug design for over 50 y. However, this model by itself fails to predict the magnitude of enzymes' rate accelerations. We construct a thermodynamic framework that begins with the classic concept of differential binding but invokes additional terms that are needed to account for subtle effects in the catalytic cycle's proton inventory. Although the model presented can be applied generally, this analysis focuses on ketosteroid isomerase (KSI) as an example, where recent experiments along with a large body of kinetic and thermodynamic data have provided strong support for the noncanonical thermodynamic contribution described. The resulting analysis precisely predicts the free energy barrier of KSI's reaction as determined from transition-state theory using only empirical thermodynamic data. This agreement is suggestive that a complete free energy inventory of the KSI catalytic cycle has been identified.enzyme catalysis | Pauling's paradigm | transition state stabilization | differential acidity E nzymes are generally viewed as devices that use protein architecture to precisely position several key functional groups and electrostatic partners with respect to the substrate (1-4). This structure-based approach is complemented by thermodynamic models, which provide abstract but quantitative descriptions of these interactions, including those that escape visual intuition. In particular, the model shown in Fig. 1, first advanced by Kurz (5), has served as a dominant paradigm for conceptualizing enzyme catalysis for 50 y. This thermodynamic cycle formalizes Pauling's hypothesis that enzymes are complementary in structure to the transition states of the reactions they catalyze (6, 7) and, combined with physical models to describe the enzyme-transition state interactions, it can help determine what leads to enzymes' catalytic effects (8).According to an elementary formulation of transition-state theory (TST), a rate constant, k, is determined by the position of a pseudoequilibrium (represented by K ‡ ) to the activated complex (9). In this depiction, chemical reactions are treated as equilibria between ground states and transition states (Fig. 1A). An enzyme's accelerated rate (k cat >> k uncat ) is represented by a relatively large value of K ‡ cat compared with K ‡ uncat (Fig. 1B). This implies that the enzyme must bind its cognate transition state very tightly, so the enzyme's dissociation constant to the transition state ðK TS D Þ is very small (2, 5). In this model, the overall rate constants are taken to be proportional to K ‡ . By solving the thermodynamic cycle of Fig. 1A, one obtainswhere k cat and k uncat are the enzyme-catalyzed and uncatalyzed unimolecular rate constants, respectively, and K S D and K TS D are dissociation constants of the enzyme to sub...

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Section: Discussionmentioning

confidence: 98%

mentioning

confidence: 99%

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Thermodynamic framework for identifying free energy inventories of enzyme catalytic cycles

Fried

Boxer

2013

Proc. Natl. Acad. Sci. U.S.A.

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“…However, sometimes the conclusion is that we have a dissociative TS based on the unjustified assumption that the distance between the nucleophile and leaving group can be used to classify a dissociative TS, or just on the belief that solution experiments point to such TS. The problem is that TSAs cannot tell us directly about the actual TS (Barbany et al 2003). In other words, the correct strategy must involve reliable computations of both the real TS and the TSA, followed by use of the calculated difference as well as the observed TSA structure as a guide for the ' observed' TS structure.…”

Section: Mechanistic Insights Into Phosphate Hydrolysis In Proteinsmentioning

confidence: 99%

Why nature really chose phosphate

Kamerlin

Sharma

Prasad

et al. 2013

Quart. Rev. Biophys.

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339

448

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Phosphoryl transfer plays key roles in signaling, energy transduction, protein synthesis, and maintaining the integrity of the genetic material. On the surface, it would appear to be a simple nucleophile displacement reaction. However, this simplicity is deceptive, as, even in aqueous solution, the low-lying d-orbitals on the phosphorus atom allow for eight distinct mechanistic possibilities, before even introducing the complexities of the enzyme catalyzed reactions. To further complicate matters, while powerful, traditional experimental techniques such as the use of linear free-energy relationships (LFER) or measuring isotope effects cannot make unique distinctions between different potential mechanisms. A quarter of a century has passed since Westheimer wrote his seminal review, ‘Why Nature Chose Phosphate’ (Science 235 (1987), 1173), and a lot has changed in the field since then. The present review revisits this biologically crucial issue, exploring both relevant enzymatic systems as well as the corresponding chemistry in aqueous solution, and demonstrating that the only way key questions in this field are likely to be resolved is through careful theoretical studies (which of course should be able to reproduce all relevant experimental data). Finally, we demonstrate that the reason that naturereallychose phosphate is due to interplay between two counteracting effects: on the one hand, phosphates are negatively charged and the resulting charge-charge repulsion with the attacking nucleophile contributes to the very high barrier for hydrolysis, making phosphate esters among the most inert compounds known. However, biology is not only about reducing the barrier to unfavorable chemical reactions. That is, the same charge-charge repulsion that makes phosphate ester hydrolysis so unfavorable also makes it possible to regulate, by exploiting the electrostatics. This means that phosphate ester hydrolysis can not only be turned on, but also be turned off, by fine tuning the electrostatic environment and the present review demonstrates numerous examples where this is the case. Without this capacity for regulation, it would be impossible to have for instance a signaling or metabolic cascade, where the action of each participant is determined by the fine-tuned activity of the previous piece in the production line. This makes phosphate esters the ideal compounds to facilitate life as we know it.

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“…Per tal de dissenyar anticossos catalítics, un mètode habitual és el de sintetitzar químicament anàlegs a l!estat de transició de la reacció objectiu i utilitzar aquests anàlegs com a haptens per tal de sintetitzar el nou anticòs catalític a través de processos immunològics [96][97][98][99][100] .…”

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Proteïnes com a microreactors en fotoquímica supramolecular

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On the Generation of Catalytic Antibodies by Transition State Analogues

Cited by 36 publications

References 50 publications

Thermodynamic framework for identifying free energy inventories of enzyme catalytic cycles

Thermodynamic framework for identifying free energy inventories of enzyme catalytic cycles

Why nature really chose phosphate

Proteïnes com a microreactors en fotoquímica supramolecular

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