Construction and Crystal Structure of Recombinant STNV Capsids
Construction and Crystal Structure of Recombinant STNV Capsids
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DOI：
10.1016/j.jmb.2011.07.062
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发表时间：
2011-10-14
影响因子：
5.6
通讯作者：
Phillips, Simon E. V.
中科院分区：
文献类型：
--
作者：
Lane, Stephen W.;Dennis, Caitriona A.;Phillips, Simon E. V.
在大肠杆菌中表达了一种密码子优化基因，以产生卫星烟草坏死病毒的外壳蛋白（CP）。这种蛋白质在体内组装成衣壳，与T=1野生型病毒的衣壳非常相似。这些病毒样颗粒（vlp）包装重组mRNA转录物，并可以在不同的缓冲条件下拆卸和重组。VLP的x射线晶体结构已在1.4埃分辨率下得到求解和细化，结果显示与野生型卫星烟草坏死病毒非常相似，除了可以去除二十面体对称约束以显示亚基之间的差异，可能是由于晶体包装。另一项低分辨率x射线晶体结构测定显示，有序的RNA片段位于衣壳内表面附近，靠近由投射到粒子内部的n端螺旋形成的基本簇。RNA由一个3-bp螺旋茎的多个拷贝组成，在3'端有一个未配对的碱基，可能由许多短的茎环组成，其中环区是无序的。RNA的排列与在其他卫星病毒中观察到的不同。(C) 2011 Elsevier Ltd.版权所有。
A codon-optimised gene has been expressed in Escherichia coli to produce the coat protein (CP) of the Satellite Tobacco Necrosis Virus. This protein assembles in vivo into capsids closely resembling those of the T=1 wildtype virus. These virus-like particles (VLPs) package the recombinant mRNA transcript and can be disassembled and reassembled using different buffer conditions. The X-ray crystal structure of the VLP has been solved and refined at 1.4 angstrom resolution and shown to be very similar to that of wild-type Satellite Tobacco Necrosis Virus, except that icosahedral symmetry constraints could be removed to reveal differences between subunits, presumably owing to crystal packing. An additional low-resolution X-ray crystal structure determination revealed well-ordered RNA fragments lodged near the inside surface of the capsid, close to basic clusters formed by the N-terminal helices that project into the interior of the particle. The RNA consists of multiple copies of a 3-bp helical stem, with a single unpaired base at the 3' end, and probably consists of a number of short stem loops where the loop region is disordered. The arrangement of the RNA is different from that observed in other satellite viruses. (C) 2011 Elsevier Ltd. All rights reserved.