Lee S, Owen K E, Choi H K, Lee H, Lu G, Wengler G, Brown D T, Rossmann M G, Kuhn R J
Department of Biological Sciences, Purdue University, West Lafayette, IN 47907-1392, USA.
Structure. 1996 May 15;4(5):531-41. doi: 10.1016/s0969-2126(96)00059-7.
Many enveloped viruses exit cells by budding from the plasma membrane. The driving force for budding is the interaction of an inner protein nucleocapsid core with transmembrane glycoprotein spikes. The molecular details of this process are ill defined. Alphaviruses, such as Sindbis virus (SINV) and Semliki Forest virus (SFV), represent some of the simplest enveloped viruses and have been well characterized by structural, genetic and biochemical techniques. Although a high-resolution structure of an alphavirus has not yet been attained, cryo-electron microscopy (cryo-EM) has been used to show the multilayer organization at 25 A resolution. In addition, atomic resolution studies are available of the C-terminal domain of the nucleocapsid protein and this has been modeled into the cryo-EM density.
A recombinant form of Sindbis virus core protein (SCP) was crystallized and found to diffract much better than protein extracted from the virus (2.0 A versus 3.0 A resolution). The new structure showed that amino acids 108 to 111 bind to a specific hydrophobic pocket in neighboring molecules. Re-examination of the structures derived from virus-extracted protein also showed this 'N-terminal arm' binding to the same hydrophobic pocked in adjacent molecules. It is proposed that the binding of these capsid residues into the hydrophobic pocket of SCP mimics the binding of E2 (one of two glycoproteins that penetrate the lipid bilayer of the viral envelope) C-terminal residues in the pocket. Mutational studies of capsid residues 108 and 110 confirm their role in capsid assembly.
Structural and mutational analyses of residues within the hydrophobic pocket suggest that budding results in a switch between two conformations of the capsid hydrophobic pocket. This is the first description of a viral budding mechanism in molecular detail.
许多包膜病毒通过从质膜出芽的方式离开细胞。出芽的驱动力是内部蛋白质核衣壳核心与跨膜糖蛋白刺突之间的相互作用。这一过程的分子细节尚不明确。甲病毒，如辛德毕斯病毒（SINV）和Semliki森林病毒（SFV），代表了一些最简单的包膜病毒，并且已经通过结构、遗传和生化技术得到了很好的表征。尽管尚未获得甲病毒的高分辨率结构，但冷冻电子显微镜（cryo-EM）已被用于以25埃分辨率显示其多层结构。此外，已有关于核衣壳蛋白C末端结构域的原子分辨率研究，并且已将其建模到冷冻电子显微镜密度图中。
一种重组形式的辛德毕斯病毒核心蛋白（SCP）被结晶，发现其衍射效果比从病毒中提取的蛋白好得多（分辨率为2.0埃，而从病毒中提取的蛋白为3.0埃）。新结构表明，氨基酸108至111与相邻分子中的一个特定疏水口袋结合。对从病毒提取的蛋白所衍生结构的重新检查也显示，这个“N末端臂”与相邻分子中的相同疏水口袋结合。有人提出，这些衣壳残基与SCP疏水口袋的结合模拟了E2（穿透病毒包膜脂质双层的两种糖蛋白之一）C末端残基在该口袋中的结合。对衣壳残基108和110的突变研究证实了它们在衣壳组装中的作用。
对疏水口袋内残基的结构和突变分析表明，出芽导致衣壳疏水口袋的两种构象之间发生转换。这是对病毒出芽机制分子细节的首次描述。
