A role for naturally occurring variation of the murine coronavirus spike protein in stabilizing association with the cellular receptor
A role for naturally occurring variation of the murine coronavirus spike protein in stabilizing association with the cellular receptor
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DOI：
10.1128/jvi.71.4.3129-3137.1997
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发表时间：
1997-04-01
影响因子：
5.4
通讯作者：
Gallagher, TM
中科院分区：
文献类型：
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作者：
Gallagher, TM
鼠肝炎病毒（MHV）是一种冠状病毒，通过从病毒体膜上突出的刺突蛋白（S）与其细胞受体（MHVR）结合而引发感染。不同MHV毒株之间S蛋白的结构差异很大，这种差异通常被认为是决定MHV感染的毒株特异性病理的重要因素，可能是通过影响MHV和MHVR之间的相互作用。为了解决S变异和受体结合之间的关系，开发了能够测量MHV和MHVR之间相互作用的测定法。该测定法利用MHVR的新可溶形式sMHVR-Ig，其包含与人免疫球蛋白G1的Fc部分融合的MHVR的病毒结合免疫球蛋白样结构域。sMHVR-Ig在人293 EBNA细胞中以二硫键连接的二聚体形式稳定表达，并通过Fc结构域固定于Sepharose-蛋白G上，所得的Sepharose珠用于吸附放射性标记的MHV颗粒，在4 ℃下，该珠特异性吸附两种原型MHV株，MHV JHM（菌株4）和MHV的组织培养适应性突变体JHM，JHMX菌株。转移至37 ° C导致JHM洗脱，但JHMX不洗脱。JHM的体外观察在37 ℃下从其受体洗脱的JHMX（但不是JHMX）被相应的37 ℃洗脱的受体相关的JHM（但不是JHMX）洗脱。受体结合维持的这种差异的基础与JHMX S蛋白中存在的大的缺失突变相关，由于sMHVR-Ig表现出相对热稳定的与包含缺失的牛痘病毒表达的S蛋白的结合，这些结果表明，冠状病毒S蛋白中自然发生的突变影响了与宿主细胞最初相互作用的稳定性，从而有助于成功感染病毒体的可能性病毒进入特征的这些变化可能导致冠状病毒具有新的致病特性。
Murine hepatitis virus (MHV), a coronavirus, initiates infection by binding to its cellular receptor (MHVR) via spike (S) proteins projecting from the virion membrane, The structures of these S proteins vary considerably among MHV strains, and this variation is generally considered to be important in determining the strain-specific pathologies of MHV infection, perhaps by affecting the interaction between MHV and the MHVR. To address the relationships between S variation and receptor binding, assays capable of measuring interactions between MHV and MHVR mere developed, The assays made use of a novel soluble form of the MHVR, sMHVR-Ig, which comprised the virus-binding immunoglobulin-like domain of MHVR fused to the Fc portion of human immunoglobulin G1. sMHVR-Ig was stably expressed as a disulfide-linked dimer in human 293 EBNA cells and was immobilized to Sepharose-protein G via the Fc domain, The resulting Sepharose beads were used to adsorb radiolabelled MHV particles, At 4 degrees C, the beads specifically adsorbed two prototype MHV strains, MHV JHM (strain 4) and a tissue culture-adapted mutant of MHV JHM, the JHMX strain, A shift to 37 degrees C resulted in elution of JHM but not JHMX. This in vitro observation of JHM (but not JHMX) elution from its receptor at 37 degrees C was paralleled by a corresponding 37 degrees C elution of receptor-associated JHM (but not JHMX) from tissue culture cells, The basis for this difference in maintenance of receptor association was correlated with a large deletion mutation present within the JHMX S protein, as sMHVR-Ig exhibited relatively thermostable binding to vaccinia virus-expressed S proteins containing the deletion, These results indicate that naturally occurring mutations in the coronavirus S protein affect the stability of the initial interaction with the host cell and thus contribute to the likelihood of successful infection by incoming virions. These changes in virus entry features may result in coronaviruses with novel pathogenic properties.