Lee B M, De Guzman R N, Turner B G, Tjandra N, Summers M F
Howard Hughes Medical Institute, University of Maryland, Baltimore County 21250, USA.
J Mol Biol. 1998 Jun 12;279(3):633-49. doi: 10.1006/jmbi.1998.1766.
The HIV-1 nucleocapsid protein (NC) contains two CCHC-type zinc knuckle domains that are essential for genome recognition, packaging and infectivity. The solution structure of the protein has been determined independently by three groups. Although the structures of the individual zinc knuckle domains are similar, two of the studies indicated that the knuckles behave as independently folded, non-interacting domains connected by a flexible tether, whereas one study revealed the presence of interknuckle NOE cross-peaks, which were interpreted in terms of a more compact structure in which the knuckles are in close proximity. We have collected multidimensional NMR data for the recombinant, isotopically labeled HIV-1 NC protein, and confirmed the presence of weak interknuckle NOEs. However, the NOE data are not consistent with a single protein conformation. 15N NMR relaxation studies reveal that the two zinc knuckle domains possess different effective rotational correlation times, indicating that the knuckles are not tumbling as a single globular domain. In addition, the 1H NMR chemical shifts of isolated zinc knuckle peptides are very similar to those of the intact protein. The combined results indicate that the interknuckle interactions, which involve the close approach of the side-chains of Phe16 and Trp37, are transitory. The solution behavior of NC may be best considered as a rapid equilibrium between conformations with weakly interacting and non-interacting knuckle domains. This inherent conformational flexibility may be functionally important, enabling adaptive binding of NC to different recognition elements within the HIV-1 psi-RNA packaging signal.
HIV-1核衣壳蛋白（NC）含有两个CCHC型锌指结构域，这对于基因组识别、包装和感染性至关重要。该蛋白的溶液结构已由三个研究小组独立确定。尽管各个锌指结构域的结构相似，但其中两项研究表明，锌指结构域表现为通过柔性连接链相连的独立折叠、非相互作用的结构域，而另一项研究则揭示了锌指间存在核Overhauser效应（NOE）交叉峰，这被解释为一种更紧凑的结构，其中锌指结构域紧密相邻。我们收集了重组的、同位素标记的HIV-1 NC蛋白的多维核磁共振（NMR）数据，并证实了锌指间存在微弱的NOE。然而，NOE数据与单一的蛋白质构象不一致。15N NMR弛豫研究表明，两个锌指结构域具有不同的有效旋转相关时间，这表明锌指结构域并非作为一个单一的球状结构域进行翻滚。此外，分离的锌指肽段的1H NMR化学位移与完整蛋白的化学位移非常相似。综合结果表明，涉及苯丙氨酸16和色氨酸37侧链紧密靠近的锌指间相互作用是短暂的。NC的溶液行为可能最好被视为具有弱相互作用和非相互作用锌指结构域的构象之间的快速平衡。这种固有的构象灵活性在功能上可能很重要，使NC能够适应性地结合HIV-1 ψ-RNA包装信号内的不同识别元件。
