Ramanujam M, Hofmann J, Nakhasi H L, Atreya C D
Center for Biologics Evaluation and Research (CBER), Food and Drug Administration, Bethesda, MD 20892, USA.
Virus Res. 2001 Dec 4;81(1-2):151-6. doi: 10.1016/s0168-1702(01)00374-4.
The role of three N-linked glycosylation sites in rubella virus (RV) E1 protein on virion release was analyzed by transfecting Vero 76 cells with infectious RV RNA (Robo302WT) containing isoleucine substitutions at N76, N177, and N209 (individually and in combinations). RV RNAs were detected and found to retain substitutions in the transfected cells, but RV capsid indicative of infection was undetectable, except for in Robo302WT and Robo302-N177I transfected cells. Only culture supernatants of Robo302WT and Robo302-N177I RNA transfected cells were positive for RV, suggestive of the virion release into the culture medium. Further, detection of intracellular RV E1 and newly released virion-associated E1 was possible only from cells previously incubated with Robo302-N177I and Robo302WT culture supernatants, suggesting that N177I substituted virus retained infectivity. These results suggest that while glycosylation at N177 is not critical, N76I and N209I mutations are lethal to RV viability.
通过用在N76、N177和N209处含有异亮氨酸替代（单独和组合）的感染性风疹病毒（RV）RNA（Robo302WT）转染Vero 76细胞，分析了风疹病毒（RV）E1蛋白上三个N-连接糖基化位点在病毒粒子释放中的作用。检测到RV RNA并发现其在转染细胞中保留了替代，但除了在Robo302WT和Robo302-N177I转染细胞中外，未检测到指示感染的RV衣壳。只有Robo302WT和Robo302-N177I RNA转染细胞的培养上清液对RV呈阳性，提示病毒粒子释放到培养基中。此外，只有从先前用Robo302-N177I和Robo302WT培养上清液孵育的细胞中才能检测到细胞内RV E1和新释放的病毒粒子相关E1，这表明N177I替代病毒保留了感染性。这些结果表明，虽然N177处的糖基化并不关键，但N76I和N209I突变对RV活力是致命的。
