Redzic Jasmina S, Duff Michael R, Blue Ashley, Pitts Todd M, Agarwal Pratul, Eisenmesser Elan Zohar
Department of Biochemistry and Molecular Genetics, School of Medicine, University of Colorado Denver, Denver, CO, United States.
Biochemistry and Cellular and Molecular Biology Department, University of Tennessee, Knoxville, TN, United States.
Front Mol Biosci. 2021 May 20;8:691208. doi: 10.3389/fmolb.2021.691208. eCollection 2021.
The biliverdin reductase B (BLVRB) class of enzymes catalyze the NADPH-dependent reduction of multiple flavin substrates and are emerging as critical players in cellular redox regulation. However, the role of dynamics and allostery have not been addressed, prompting studies here that have revealed a position 15 Å away from the active site within human BLVRB (T164) that is inherently dynamic and can be mutated to control global micro-millisecond motions and function. By comparing the inherent dynamics through nuclear magnetic resonance (NMR) relaxation approaches of evolutionarily distinct BLVRB homologues and by applying our previously developed Relaxation And Single Site Multiple Mutations (RASSMM) approach that monitors both the functional and dynamic effects of multiple mutations to the single T164 site, we have discovered that the most dramatic mutagenic effects coincide with evolutionary changes and these modulate coenzyme binding. Thus, evolutionarily changing sites distal to the active site serve as dynamic "dials" to globally modulate motions and function. Despite the distal dynamic and functional coupling modulated by this site, micro-millisecond motions span an order of magnitude in their apparent kinetic rates of motions. Thus, global dynamics within BLVRB are a collection of partially coupled motions tied to catalytic function.
胆绿素还原酶B（BLVRB）类酶催化多种黄素底物的NADPH依赖性还原反应，正逐渐成为细胞氧化还原调节中的关键因子。然而，动力学和别构作用的角色尚未得到研究，促使我们在此开展的研究揭示了人BLVRB（T164）中距活性位点15埃处的一个位置，该位置具有内在动力学特性，可通过突变来控制整体微毫秒级运动和功能。通过比较进化上不同的BLVRB同源物的固有动力学（采用核磁共振（NMR）弛豫方法），并应用我们先前开发的弛豫与单一位点多重突变（RASSMM）方法（该方法监测多个突变对单个T164位点的功能和动力学影响），我们发现最显著的诱变效应与进化变化一致，且这些变化调节辅酶结合。因此，活性位点远端的进化变化位点充当动态“刻度盘”，以全局调节运动和功能。尽管该位点调节了远端动力学和功能耦合，但微毫秒级运动在其明显的运动动力学速率上跨越了一个数量级。因此，BLVRB内的整体动力学是与催化功能相关的部分耦合运动的集合。
