Moseley H N, Curto E V, Krishna N R
Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham 35294-2041, USA.
J Magn Reson B. 1995 Sep;108(3):243-61. doi: 10.1006/jmrb.1995.1129.
A very general procedure entitled complete relaxation and conformational exchange matrix (CORCEMA) analysis has been developed to analyze the 2D-NOESY spectra of interacting systems undergoing multistate conformational exchange. This is an extension of earlier work from this laboratory on the methodological treatment of multistate conformational exchange [Krishna et al., Biopolymers 19, 2003 (1980)] and the theory of transferred NOESY for finite exchange off-rates [Lee and Krishna, J. Magn. Reson. 98, 36 (1992)]. The current theory is based on generalized rate matrices for relaxation and conformational exchange. The CORCEMA algorithm explicitly incorporates intermolecular dipolar cross relaxation between the molecules when they are complexed. It permits an analysis of NOESY intensities for the intra- as well as intermolecular contacts between the interacting molecules under a variety of binding conditions. Its application is illustrated on two examples of transferred NOESY simulations: (1) a two-state system involving a ligand and an enzyme forming a ligand-enzyme complex, and (2) a three-state system in which the ligand-enzyme complex can undergo a conformational transition from an "open state" to a "closed state," and can include conformational changes in both the complexed ligand and the complexed enzyme, such as hinge-bending motions. Simplifying expressions for generalized matrix analyses are derived for three limiting cases of the three-state system. This three-state example is illustrated using a hypothetical model of the hinge-bending motion in a thermolysin-inhibitor complex. It is shown that: (1) The neglect of cross relaxation between the interacting species in their complexed forms can lead to misleading conclusions on the "bound" conformation of the ligand. (2) If protein-mediated spin diffusion is dominant, caution is needed in analyses based on initial slopes alone due to one's inability to identify the exact range of the initial growth curve under poor signal/noise situations. (3) The neglect of conformational changes upon complexation, e.g., hinge-bending motions of the ligand-enzyme complex, can lead to erroneous results on the nature of "bound" conformations of the ligand. In this case, attempts to analyze the transferred NOESY data with a two-state model will result in a "virtual" conformation for the bound ligand. (4) When the hinge-bending rate is slower than the cross relaxation and enzyme off-rates, the bound conformation of a ligand deduced from the transferred NOESY experiment is more likely to represent nonspecific or weak binding in an open state of the enzyme.(ABSTRACT TRUNCATED AT 400 WORDS)
已经开发出一种名为完全弛豫和构象交换矩阵（CORCEMA）分析的通用程序，用于分析经历多态构象交换的相互作用系统的二维NOESY谱。这是本实验室早期关于多态构象交换方法处理的工作[克里希纳等人，《生物聚合物》19, 2003 (1980)]以及有限交换速率下转移NOESY理论[李和克里希纳，《磁共振杂志》98, 36 (1992)]的扩展。当前理论基于弛豫和构象交换的广义速率矩阵。CORCEMA算法明确纳入了分子形成复合物时分子间的偶极交叉弛豫。它允许在各种结合条件下分析相互作用分子之间分子内以及分子间接触的NOESY强度。通过两个转移NOESY模拟示例说明了其应用：（1）一个涉及配体和酶形成配体 - 酶复合物的双态系统，以及（2）一个三态系统，其中配体 - 酶复合物可经历从“开放状态”到“封闭状态”的构象转变，并且可以包括复合配体和复合酶中的构象变化，如铰链弯曲运动。针对三态系统的三种极限情况推导了广义矩阵分析的简化表达式。使用嗜热菌蛋白酶 - 抑制剂复合物中铰链弯曲运动的假设模型说明了这个三态示例。结果表明：（1）忽略相互作用物种在其复合形式下的交叉弛豫可能会导致关于配体“结合”构象的误导性结论。（2）如果蛋白质介导的自旋扩散占主导，由于在信号/噪声较差的情况下无法确定初始增长曲线的确切范围，仅基于初始斜率进行分析时需要谨慎。（3）忽略复合时的构象变化，例如配体 - 酶复合物的铰链弯曲运动，可能会导致关于配体“结合”构象性质的错误结果。在这种情况下，尝试用双态模型分析转移NOESY数据将导致结合配体的“虚拟”构象。（4）当铰链弯曲速率慢于交叉弛豫和酶解离速率时，从转移NOESY实验推导的配体结合构象更可能代表酶开放状态下的非特异性或弱结合。（摘要截断于400字）
