Structural investigation of reaction intermediates in glycolytic aldolases

关于酶反应的催化作用和热力学，已经获得了大量的信息。然而，对酶中构象过程的动力学和能量学知之甚少。酶如何通过催化循环来处理它们的反应中间体仍然是酶学中的一个未解决的问题。挑战在于解开蛋白质运动和酶功能之间的密切联系。在室温（RT）下进行的蛋白质晶体学是一种可用于分析酶构象运动的工具。我们将根据蛋白质序列中每个氨基酸（aa）的一个或多个构象状态，分析从晶体学研究中获得的电子密度。如果一个aa构象与邻近的aa构象重叠，则它可以代表传递酶中构象运动的管道。这些网络可以延伸到整个酶，并对应于不改变原子平衡位置的动力学驱动的变构。我们的目标是辨别哪些接触网络与酶反应中间体相互作用，以了解这些相互作用如何促进一种中间体向另一种中间体的化学转化。我们已经广泛地研究了糖酵解酶醛缩酶，以解开活性位点与反应中间体的相互作用，现在希望了解构象运动如何通过催化循环的反应中间体的进展。醛缩酶是负责产生储存的生物能量和代谢前体的酶。

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.