Smith C C, Aurelian L
Department of Pharmacology and Experimental Therapeutics, University of Maryland School of Medicine, Baltimore 21201, USA.
Virology. 1997 Aug 4;234(2):235-42. doi: 10.1006/viro.1997.8645.
The large subunit of herpes simplex virus type 2 (HSV-2) ribonucleotide reductase (ICP10) was identified in sucrose gradient-purified HSV-2 virions by immunoprecipitation/immunoblotting with antibody specific for the protein kinase (PK) domain. Immunoblotting of individual gradient fractions indicated that ICP10 cosediments with the major capsid protein and the highest virus titers. ICP10 was not labeled by iodination of purified virions, indicating that it is not located on the virion surface. After envelope glycoproteins were removed by detergent treatment, ICP10 was associated with capsid-tegument particles and became sensitive to trypsin digestion. The capsid-tegument-associated ICP10 was phosphorylated and had PK activity in vitro and on Immobilon membranes. A mutant ICP10 protein deleted in the PK domain (p95) was also associated with purified virions (ICP10deltaPK virus) but it lacked PK activity. The data indicate that ICP10 is contained within the tegument component where it retains intrinsic PK activity.
通过用针对蛋白激酶(PK)结构域的特异性抗体进行免疫沉淀/免疫印迹,在蔗糖梯度纯化的单纯疱疹病毒2型(HSV-2)病毒粒子中鉴定出了HSV-2核糖核苷酸还原酶的大亚基(ICP10)。对各个梯度级分进行免疫印迹表明,ICP10与主要衣壳蛋白及最高病毒滴度共沉降。纯化病毒粒子的碘化标记未显示ICP10被标记,这表明它不在病毒粒子表面。经去污剂处理去除包膜糖蛋白后,ICP10与衣壳-内膜颗粒相关联,并变得对胰蛋白酶消化敏感。与衣壳-内膜相关的ICP10被磷酸化,并且在体外和在Immobilon膜上具有PK活性。在PK结构域缺失的突变型ICP10蛋白(p95)也与纯化的病毒粒子相关联(ICP10deltaPK病毒),但它缺乏PK活性。数据表明,ICP10包含在内膜成分中,在那里它保留了内在的PK活性。