Stiller John B, Kerns S Jordan, Hoemberger Marc, Cho Young-Jin, Otten Renee, Hagan Michael F, Kern Dorothee
Department of Biochemistry and Howard Hughes Medical Institute, Brandeis University, Waltham, Massachusetts 02452, United States.
Present addresses: S.J.K. 27 Drydock Ave, Boston MA 02110 , M.H. 225 Binney St, Cambridge, MA 02142, Y.J.C. 733 Concord Ave Cambridge, MA 02138.
Nat Catal. 2019 Aug;2(8):726-734. doi: 10.1038/s41929-019-0307-6. Epub 2019 Jun 24.
Protein conformational changes are frequently essential for enzyme catalysis, and in several cases, shown to be the limiting factor for overall catalytic speed. However, a structural understanding of corresponding transition states, needed to rationalize the kinetics, remains obscure due to their fleeting nature. Here, we determine the transition-state ensemble of the rate-limiting conformational transition in the enzyme adenylate kinase, by a synergistic approach between experimental high-pressure NMR relaxation during catalysis and molecular dynamics simulations. By comparing homologous kinases evolved under ambient or high pressure in the deep-sea, we detail transition state ensembles that differ in solvation as directly measured by the pressure dependence of catalysis. Capturing transition-state ensembles begins to complete the catalytic energy landscape that is generally characterized by structures of all intermediates and frequencies of transitions among them.
蛋白质构象变化对于酶催化通常至关重要，并且在一些情况下，已被证明是整体催化速度的限制因素。然而，由于相应过渡态转瞬即逝的性质，为使动力学合理化而需要的对其结构的理解仍然模糊不清。在这里，我们通过催化过程中实验性高压核磁共振弛豫与分子动力学模拟之间的协同方法，确定了腺苷酸激酶中限速构象转变的过渡态系综。通过比较在深海环境压力或高压下进化的同源激酶，我们详细阐述了通过催化作用的压力依赖性直接测量的、在溶剂化方面存在差异的过渡态系综。捕捉过渡态系综开始完善通常由所有中间体的结构及其之间转变频率所表征的催化能量景观。
