Yu I-M, Holdaway H A, Chipman P R, Kuhn R J, Rossmann M G, Chen J
Department of Biological Sciences, Purdue University, West Lafayette, Indiana 47907, USA.
J Virol. 2009 Dec;83(23):12101-7. doi: 10.1128/JVI.01637-09. Epub 2009 Sep 16.
Flavivirus assembles into an inert particle that requires proteolytic activation by furin to enable transmission to other hosts. We previously showed that immature virus undergoes a conformational change at low pH that renders it accessible to furin (I. M. Yu, W. Zhang, H. A. Holdaway, L. Li, V. A. Kostyuchenko, P. R. Chipman, R. J. Kuhn, M. G. Rossmann, and J. Chen, Science 319:1834-1837, 2008). Here we show, using cryoelectron microscopy, that the structure of immature dengue virus at pH 6.0 is essentially the same before and after the cleavage of prM. The structure shows that after cleavage, the proteolytic product pr remains associated with the virion at acidic pH, and that furin cleavage by itself does not induce any major conformational changes. We also show by liposome cofloatation experiments that pr retention prevents membrane insertion, suggesting that pr is present on the virion in the trans-Golgi network to protect the progeny virus from fusion within the host cell.
黄病毒组装成一种无活性的颗粒，需要弗林蛋白酶进行蛋白水解激活才能传播到其他宿主。我们之前表明，未成熟病毒在低pH值下会发生构象变化，使其能够被弗林蛋白酶作用（I.M. Yu、W. Zhang、H.A. Holdaway、L. Li、V.A. Kostyuchenko、P.R. Chipman、R.J. Kuhn、M.G. Rossmann和J. Chen，《科学》319:1834 - 1837，2008）。在此我们利用冷冻电子显微镜表明，在pH 6.0条件下，未成熟登革病毒在prM裂解前后的结构基本相同。该结构表明，裂解后，蛋白水解产物pr在酸性pH值下仍与病毒粒子结合，并且弗林蛋白酶裂解本身不会诱导任何主要的构象变化。我们还通过脂质体共漂浮实验表明，pr的保留可防止膜插入，这表明pr存在于反式高尔基体网络中的病毒粒子上，以保护子代病毒在宿主细胞内不发生融合。
