摘要
乙肝核心抗原(HBcAg)蛋白基因(C基因)在酿酒酵母中表达,表达产物经过分离和Sepharose CL-4B柱子的初步纯化。产物经SDS-PAGE和Western blotting鉴定为一分子量约21.5kDa的多肽。再经蔗糖密度梯度超离心和CsCl等密度梯度超离心等过程而被纯化。分管收集的超离心纯化产物经ELISA抗原活性检测和密度分析,可知ELISA反应强度较高的收集管中的颗粒密度主要分布在1.27g/mL和1.40 g/mL两个峰值处。将rHBcAg抗原活性最高的收集管合并,再经TEM观察,发现酵母表达的rHBcAg蛋白(核心蛋白)能自主装配成大小不同的两种核心颗粒,大颗粒直径约为30.1±2.4 nm,小颗粒直径约为21.5±3.3 nm。这表明,酿酒酵母表达的rHBcAg颗粒具有大小不同的二态性,其生物学意义还未明了,需进一步研究和探讨。
Hepatitis B virus core antigen protein gene (C gene) was expressed in Saccharomyces cerevisiae and the products (rHBcAg particles) were purified from crude lysate of the yeast by three steps: Sephrose CL-4B chromatography, sucrose step-gradient ultracentrifugation, and CsCl-isopycnic ultracentrifugation. rHBcAg was synthesized in yeast cells as a particle consisting of polypeptides which have a molecular weight of 21.5 kDa (p21.5). ELISA and CsCl- isopycnic ultracentrifugation purification products (rHBcAg particles) in each fraction indicated that the particle densities of the fraction with higher rHBcAg antigenicity mainly distributed at the densities of 1.27g·mL-1 and 1.40 g·mL-1, respectively. Observation of the purified products (rHBcAg particles) by TEM indicated that rHBcAg peptides could self-assemble into two kinds of different sizes of rHBcAg particles (core particles); the large particle was about 30.1±2.4 nm in diameter and the small particle about 21.5±3.3 nm. This indicated that the rHBcAg particles displayed dimorphism, but the biological significance of this dimorphism is not clear and needs to be studied further.
出处
《中国病毒学》
CSCD
2004年第2期101-104,共4页
Virologica Sinica
