Comparative analysis of functional domains and motifs within glycoprotein gH of Epstein-Barr Virus and Pseudorabies Virus, and their functional role in the herpesviral fusion process

疱疹病毒是最复杂的病毒之一，疱疹病毒粒子由40多种蛋白质组成。根据生物学特性和序列比较，疱疹病毒科分为三个亚科：α-、β-和伽马-疱疹病毒亚科。宿主细胞的感染在附着后开始，要么是通过病毒被膜与质膜直接融合，要么是在内吞之后与内体膜融合。虽然许多病毒的膜融合是由一种或两种病毒蛋白完成的，但疱疹病毒的进入需要四种不同蛋白的协同作用。渗透的第一步是将亚家族特异性病毒蛋白与宿主细胞受体结合。稳定的结合触发融合，而融合依赖于由糖蛋白(G)B和Gh/gl组成的保守的核心融合机制。缺乏这两种蛋白质中的任何一种的突变体都无法进入细胞。GB的晶体结构显示出III类融合蛋白的特征，然而，GB不足以介导有效的融合，而需要异二聚体Gh/Gl复合体。Gh和gl在融合过程中的作用仍然是个谜。最近解决的单纯疱疹病毒2型和EB病毒(EBV)的Gh/g1以及伪狂犬病病毒(PRV)Gh的核心区的晶体结构表明，Gh与四个明显可分离的Gh区具有惊人的相似性。该建议旨在比较PRV和EBV Gh内的功能结构域和基序，以及它们在融合过程中的作用。为此，不同的Gh结构域将在y-疱疹病毒成员PRV的Gh和属于y-疱疹病毒亚科的EBV的Gh之间交换，识别的基序以及关键氨基酸将发生突变。嵌合体和突变的Gh分子将在融合试验中进行测试，以确定与gl和gb的相互作用，以及相应缺失突变体的互补性。这些结果将有助于理解融合过程，并可能导致新的抗病毒策略。

Herpesviruses are amongst the most complex viruses and herpes virions consist of more than 40 proteins. Three subfamilies, alpha-, beta-, and gamma-Herpesvirinae, have been classified within the Herpesviridae based on biological properties and sequence comparison. Infection of host cells starts after attachment either by direct fusion of the viral envelope with the plasma membrane or, after endo-cytic uptake, with the endosomal membrane. While membrane fusion in many viruses is accomplished by one or two viral proteins, herpesvirus entry requires the concerted action of four different proteins. The first step in penetration is binding of subfamily-specific viral proteins to host cell receptors. Stable binding triggers fusion which relies on a conserved core fusion machinery consisting of glycoproteins (g)B and gH/gL. Mutants lacking either of these proteins are unable to enter cells. The crystal structure of gB shows signatures of class III fusion proteins, however, gB is not sufficient to mediate efficient fusion but requires the heterodimeric gH/gL complex. The role of gH and gL in the fusion process is still enigmatic. The recently solved crystal structures for gH/gL of Herpes Simplex Virus-2 and Epstein-Barr virus (EBV), as well as of a core domain of Pseudorabies virus (PrV) gH revealed surprising similarity with four clearly separable gH domains. This proposal aims at the comparison of functional domains and motifs within gH of PrV and EBV, and their role in the fusion process. To this end, the different gH domains will be swapped between gH of PrV, a member of the y-herpesviruses and of EBV, which belongs to the y-Herpesvirinae, and identified motifs as well as key amino acids will be mutated. The chimeras and mutated gH molecules will be tested in fusion assays, for interaction with gL and gB, and for complementation of corresponding deletion mutants. These results will help to understand the fusion process and could lead to new antiviral strategies.