Ennifar E, Yusupov M, Walter P, Marquet R, Ehresmann B, Ehresmann C, Dumas P
UPR9002 - Institut de Biologie Moléculaire et Cellulaire du CNRS, Strasbourg cedex, F-67084, France.
Structure. 1999 Nov 15;7(11):1439-49. doi: 10.1016/s0969-2126(00)80033-7.
An important step in retroviral replication is dimerization of the genomic RNA prior to encapsidation. Dimerization is initiated by the formation of a transient 'kissing-loop complex' that is thought to be subsequently matured into an extended duplex by the nucleocapsid protein (NCp). Although chemical probing and nuclear magnetic resonance spectroscopy have provided insight into the structure of the kissing-loop structure, no structural information concerning the extended-duplex state is available so far.
The structure of a minimal HIV-1 RNA dimerization initiation site has been solved at 2.3 A resolution in two different space groups. It reveals a 22 base pair extended duplex with two noncanonical Watson-Crick-like G-A mismatches, each adjacent to a bulged-out adenine. The structure shows significant asymmetry in deep groove width and G-A base-pair conformations. A network of eight magnesium cations was clearly identified, one being unusually chelated by the 3' phosphate of each bulge across an extremely narrowed deep major groove.
These crystal structures represent the putative matured form of the initial kissing-loop complex. They show the ability of this self-complementary RNA hairpin loop to acquire a more stable extended duplex structure. Both bulged adenines form a striking 'base grip' that could be a recognition signal, either in cis for another viral RNA sequence, or in trans for a protein, possibly the NCp. Magnesium binding might be important to promote and stabilize the observed extrahelical conformation of these bulges.
逆转录病毒复制过程中的一个重要步骤是基因组RNA在衣壳化之前发生二聚化。二聚化由短暂的“吻式环复合物”的形成引发，该复合物随后被认为通过核衣壳蛋白（NCp）成熟为延伸双链体。尽管化学探针和核磁共振光谱已为吻式环结构的结构提供了见解，但迄今为止尚无关于延伸双链体状态的结构信息。
在两种不同的空间群中以2.3埃的分辨率解析了最小HIV-1 RNA二聚化起始位点的结构。它揭示了一个22个碱基对的延伸双链体，带有两个非经典的类似沃森-克里克的G-A错配，每个错配都与一个凸出的腺嘌呤相邻。该结构在深沟宽度和G-A碱基对构象上显示出明显的不对称性。清楚地鉴定出由八个镁阳离子组成的网络，其中一个被每个凸起的3'磷酸异常螯合，穿过极其狭窄的深主沟。
这些晶体结构代表了初始吻式环复合物的假定成熟形式。它们展示了这种自我互补RNA发夹环获得更稳定的延伸双链体结构的能力。两个凸出的腺嘌呤形成了一个引人注目的“碱基抓手”，这可能是一种识别信号，要么顺式作用于另一个病毒RNA序列，要么反式作用于一种蛋白质，可能是核衣壳蛋白。镁结合可能对促进和稳定观察到的这些凸起的螺旋外构象很重要。
