Matusick-Kumar L, Newcomb W W, Brown J C, McCann P J, Hurlburt W, Weinheimer S P, Gao M
Department of Virology, Bristol-Myers Squibb Pharmaceutical Research Institute, Wallingford, Connecticut 06492-7660, USA.
J Virol. 1995 Jul;69(7):4347-56. doi: 10.1128/JVI.69.7.4347-4356.1995.
The herpes simplex virus type 1 protease and its substrate, ICP35, are involved in the assembly of viral capsids. Both proteins are encoded by a single open reading frame from overlapping mRNAs. The protease is autoproteolytically processed at two sites. The protease cleaves itself at the C-terminal site (maturation site) and also cleaves ICP35 at an identical site, releasing a 25-amino-acid (aa) peptide from each protein. To determine whether these 25 aa play a role in capsid assembly, we constructed a mutant virus expressing only Prb, the protease without the C-terminal 25 aa. Phenotypic analysis of the Prb virus in the presence and absence of ICP35 shows the following: (i) Prb retains the functional activity of the wild-type protease which supports virus growth in the presence of ICP35; (ii) in contrast to the ICP35 null mutant delta ICP35 virus, the Prb virus fails to grow in the absence of ICP35; and (iii) trans-complementation experiments indicated that full-length ICP35 (ICP35 c,d), but not the cleaved form (ICP35 e,f), complements the growth of the Prb virus. The most striking phenotype of the Prb virus is that only unsealed aberrant capsid structures are observed by electron microscopy in mutant-infected Vero cells. Our results demonstrate that the growth of herpes simplex virus type 1 requires the C-terminal 25 aa of either the protease or its substrate, ICP35, and that the C-terminal 25 aa are involved in the formation of sealed capsids.
单纯疱疹病毒1型蛋白酶及其底物ICP35参与病毒衣壳的组装。这两种蛋白由来自重叠mRNA的单个开放阅读框编码。该蛋白酶在两个位点进行自我蛋白水解加工。蛋白酶在C末端位点（成熟位点）切割自身，也在相同位点切割ICP35，从每种蛋白上释放出一个25个氨基酸（aa）的肽段。为了确定这25个氨基酸是否在衣壳组装中起作用，我们构建了一种仅表达Prb的突变病毒，Prb是不含C末端25个氨基酸的蛋白酶。对存在和不存在ICP35时Prb病毒的表型分析显示如下：（i）Prb保留了野生型蛋白酶的功能活性，在存在ICP35时支持病毒生长；（ii）与ICP35缺失突变体ΔICP35病毒相反，Prb病毒在不存在ICP35时无法生长；（iii）反式互补实验表明，全长ICP35（ICP35 c,d），而不是切割形式（ICP35 e,f），可互补Prb病毒的生长。Prb病毒最显著的表型是，在突变体感染的Vero细胞中，通过电子显微镜仅观察到未封闭的异常衣壳结构。我们的结果表明，单纯疱疹病毒1型的生长需要蛋白酶或其底物ICP35的C末端25个氨基酸，并且C末端25个氨基酸参与封闭衣壳的形成。
