Yamasaki K, Saito M, Oobatake M, Kanaya S
Protein Engineering Research Institute, Osaka, Japan.
Biochemistry. 1995 May 23;34(20):6587-601. doi: 10.1021/bi00020a003.
The backbone dynamics of Escherichia coli ribonuclease HI (RNase HI) in the picosecond to nanosecond time scale were characterized by a combination of measurements of 15N-NMR relaxation (T1, T2, and NOE), analyzed by a model-free approach, and molecular dynamics (MD) simulation in water. The MD simulations in water were carried out with long-range Coulomb interactions to avoid the artificial fluctuation caused by the cutoff approximation. The model-free analysis of the 15N-NMR relaxation indicated that RNase HI has a rotational correlation time of 10.9 ns at 27 degrees C. The generalized order parameter (S2) for the internal motions varied from 0.15 to 1.0, with an average value of 0.85, which is much larger than that of the RNase H domain of HIV-1 reverse transcriptase (0.78). Large internal motions (small order parameters) were observed in the N-terminal region (Leu2-Lys3), the loop between beta-strands A and B (Cys13-Gly15), the turn between alpha-helix I and beta-strand D (Glu61, His62), the loop between beta-strand D and alpha-helix II (Asp70-Tyr71), the loop between alpha-helices III and IV (Ala93-Lys96), the loop between beta-strand E and alpha-helix V (Gly123-His127), and the C-terminal region (Gln152-Val155). The effective correlation time observed in these regions varied from 0.45 ns (Glu61, Lys96) to 2.2 ns (Leu14). The order parameters calculated from the MD agreed well with those from the NMR experiment, with a few exceptions. The distributions of most of the backbone N-H vectors obtained by MD are approximately consistent with the diffusion-in-a-cone model. These distributions, however, were elliptic, with a long axis perpendicular to the plane defined by the N-H and N-C alpha vectors. Distributions supporting the axial fluctuation model or the jump-between-two-cones model were also observed in the MD simulation.
通过结合15N-NMR弛豫测量（T1、T2和NOE），采用无模型方法进行分析，并在水中进行分子动力学（MD）模拟，对大肠杆菌核糖核酸酶HI（RNase HI）在皮秒到纳秒时间尺度上的主链动力学进行了表征。在水中进行的MD模拟采用了长程库仑相互作用，以避免由截断近似引起的人为波动。对15N-NMR弛豫的无模型分析表明，RNase HI在27℃时的旋转相关时间为10.9纳秒。内部运动的广义序参数（S2）在0.15至1.0之间变化，平均值为0.85，远大于HIV-1逆转录酶的RNase H结构域（0.78）。在N端区域（Leu2-Lys3）、β链A和B之间的环（Cys13-Gly15）、α螺旋I和β链D之间的转角（Glu61、His62）、β链D和α螺旋II之间的环（Asp70-Tyr71）、α螺旋III和IV之间的环（Ala93-Lys96）、β链E和α螺旋V之间的环（Gly123-His127）以及C端区域（Gln152-Val155）观察到较大的内部运动（小序参数）。在这些区域观察到的有效相关时间从0.45纳秒（Glu61、Lys96）到2.2纳秒（Leu14）不等。从MD计算得到的序参数与NMR实验得到的数据基本一致，但有少数例外。通过MD获得的大多数主链N-H向量的分布大致符合圆锥内扩散模型。然而，这些分布是椭圆形的，长轴垂直于由N-H和N-Cα向量定义的平面。在MD模拟中也观察到了支持轴向波动模型或双锥间跳跃模型的分布。
