Structural investigation of reaction intermediates in glycolytic aldolases

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A wealth of information has been obtained about catalysis and thermodynamics of enzyme reactions. However, much less is known about the kinetics and energetics of conformational processes in the enzyme. How enzymes progress their reaction intermediates through the catalytic cycle remains an open question in enzymology. The challenge is to unravel the intimate linkage between protein motion and enzymatic function. Protein crystallography conducted at room temperature (RT) is a tool that can be used to analyze conformational motions of enzymes. We will analyze the electron density obtained from crystallographic studies in terms of one or more conformational states for each amino acid (aa) in the protein sequence. If one aa conformation overlaps with a neighboring aa conformation, it can represent a conduit for transmitting conformational motion in the enzyme. These networks can extend across the entire enzyme and correspond to dynamics-driven allostery that does not change the equilibrium positions of atoms. The goal will be to discern which contact networks interact with the enzyme reaction intermediates to understand how these interactions facilitate the chemical transformation of one intermediate into another. We have extensively investigated the glycolytic enzyme aldolase to unravel active site interactions with reaction intermediates and now wish to understand how conformational motions progress the reaction intermediates through the catalytic cycle. Aldolases are enzymes responsible for the generation of stored biological energy and metabolic precursors.