用于结构测定的单纯疱疹病毒1型糖蛋白的表达、纯化及结晶
Expression, Purification, and Crystallization of HSV-1 Glycoproteins for Structure Determination.
作者信息
White Ellen M, Stampfer Samuel D, Heldwein Ekaterina E
机构信息
Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, MA, USA.
出版信息
Methods Mol Biol. 2020;2060:377-393. doi: 10.1007/978-1-4939-9814-2_23.
Herpes simplex viruses utilize glycoproteins displayed on the viral envelope to perform a variety of functions in the viral infectious cycle. Structural and functional studies of these viral glycoproteins can benefit from biochemical, biophysical, and structural analysis of purified proteins. Here, we describe a general protocol for expression and purification of viral glycoproteins from insect cells based on those developed for the HSV-1 gB and HSV-2 gH/gL ectodomains as well as the protocol for crystallization of these glycoproteins. This protocol can be used for generating milligram amounts of wild-type (WT) or mutant gB and gH/gL ectodomains or can be adapted to produce purified ectodomains of glycoproteins from HSV or other herpesviruses for biochemical and structural studies.
单纯疱疹病毒利用病毒包膜上展示的糖蛋白在病毒感染周期中执行多种功能。对这些病毒糖蛋白的结构和功能研究可受益于对纯化蛋白的生化、生物物理和结构分析。在此，我们基于针对单纯疱疹病毒1型糖蛋白B（HSV-1 gB）和单纯疱疹病毒2型糖蛋白H/糖蛋白L（HSV-2 gH/gL）胞外域所开发的方法，描述了一种从昆虫细胞中表达和纯化病毒糖蛋白的通用方案，以及这些糖蛋白的结晶方案。该方案可用于生成毫克量的野生型（WT）或突变型gB和gH/gL胞外域，也可进行调整以生产来自单纯疱疹病毒或其他疱疹病毒的纯化糖蛋白胞外域，用于生化和结构研究。
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