Tjandra N, Wingfield P, Stahl S, Bax A
Laboratory of Chemical Physics, National Institutes of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0250, USA.
J Biomol NMR. 1996 Oct;8(3):273-84. doi: 10.1007/BF00410326.
15N NMR relaxation times in perdeuterated HIV-1 protease, complexed with the sub-nanomolar inhibitor DMP323, have been measured at 600 and 360 MHz 1H frequency. The relative magnitudes of the principal components of the inertia tensor, calculated from the X-ray coordinates of the protein-drug complex, are 1.0:0.85:0.44. The relation between the T1/T2 ratios observed for the individual backbone amides and their N-H orientation within the 3D structure of the protease dimer yields a rotational diffusion tensor oriented nearly collinear to the inertia tensor. The relative magnitudes of its principal components (1.00:1.11:1.42) are also in good agreement with hydrodynamic modeling results. The orientation and magnitude of the diffusion tensors derived from relaxation data obtained at 360 and 600 MHz are nearly identical. The anisotropic nature of the rotational diffusion has little influence on the order parameters derived from the 15N T1 and T2 relaxation times; however, if anisotropy is ignored, this can result in erroneous identification of either exchange broadening or internal motions on a nanosecond time scale. The average ratio of the T1 values measured at 360 and 600 MHz is 0.50 +/- 0.015, which is slightly larger than the value of 0.466 expected for an isotropic rigid rotor with tau c = 10.7 ns. The average ratio of the T2 values measured at 360 and 600 MHz is 1.14 +/- 0.04, which is also slightly larger than the expected ratio of 1.11. This magnetic field dependence of the T1 and T2 relaxation times suggests that the spectral density contribution from fast internal motions is not negligible, and that the chemical shift anisotropy of peptide backbone amides, on average, is larger than the 160 ppm value commonly used in 15N relaxation studies of proteins.
已在600和360 MHz的1H频率下测量了与亚纳摩尔抑制剂DMP323复合的全氘代HIV-1蛋白酶中的15N NMR弛豫时间。根据蛋白质-药物复合物的X射线坐标计算出的惯性张量主成分的相对大小为1.0:0.85:0.44。在蛋白酶二聚体的三维结构中观察到的各个主链酰胺的T1/T2比值与其N-H取向之间的关系产生了一个与惯性张量几乎共线的旋转扩散张量。其主成分的相对大小(1.00:1.11:1.42)也与流体动力学建模结果高度吻合。从360和600 MHz获得的弛豫数据得出的扩散张量的取向和大小几乎相同。旋转扩散的各向异性性质对从15N T1和T2弛豫时间得出的序参量影响很小;然而,如果忽略各向异性,这可能导致错误识别纳秒时间尺度上的交换展宽或内部运动。在360和600 MHz下测量的T1值的平均比值为0.50±0.015,略大于τc = 10.7 ns的各向同性刚性转子预期的0.466值。在360和600 MHz下测量的T2值的平均比值为1.14±0.04,也略大于预期比值1.11。T1和T2弛豫时间的这种磁场依赖性表明,快速内部运动对光谱密度的贡献不可忽略,并且肽主链酰胺的化学位移各向异性平均大于蛋白质15N弛豫研究中常用的160 ppm值。