Gas-phase electrophoretic molecular mobility analysis of size and stoichiometry of complexes of a common cold virus with antibody and soluble receptor molecules
Gas-phase electrophoretic molecular mobility analysis of size and stoichiometry of complexes of a common cold virus with antibody and soluble receptor molecules
复制标题
DOI：
10.1021/ac702463z
复制
发表时间：
2008-03-15
影响因子：
7.4
通讯作者：
Allmaier, Guenter
中科院分区：
文献类型：
--
作者：
Laschobert, Christian;Wruss, Juergen;Allmaier, Guenter
用纳米电喷雾电离气相电泳分子迁移率分析仪（nESI-GEMMA）研究了非聚集单克隆抗体和可溶性重组受体分子与二十面体无包膜人鼻病毒血清2型的附着。通过nESI-GEMMA测定的病毒质量在仪器精度（+/- 6%）范围内，接近理论值（8 × 10(6) Da），理论值是由一个病毒颗粒的所有成分（四种病毒衣壳蛋白各60个拷贝、RNA基因组和RNA连接蛋白VpG的一个拷贝）的总和计算得出的。监测不同化学计量的病毒抗体复合物（最高质量为12.5 x 10(6) Da，对应30个附着抗体）和病毒受体复合物（最高质量为8.8 x 10(6) Da，对应12个附着受体分子）的形成。通过电泳迁移率直径（EMD）得到的体积，计算了HRV配合物的化学计量。EMD的精度在+/- 0.5 nm以内，对应于相应复合物中+/- 4抗体和+/- 5受体分子的精度。在这里，我们首次展示了在常压条件下使用nESI-GEMMA实时分析高阶配合物中生物分子的大小和化学计量学。
Attachment of a nonaggregating monoclonal antibody and of a soluble recombinant receptor molecule to the icosahedral nonenveloped human rhinovirus serotype 2 was studied with a nanoelectrospray ionization gas-phase electrophoretic molecular mobility analyzer (nESI-GEMMA). The virus mass, as determined via nESI-GEMMA, was within instrument accuracy (+/- 6%) close to the theoretical value (8 x 10(6) Da) calculated from the sum of all constituents of one virus particle (60 copies of each of the four viral capsid proteins, the RNA genome, and one copy of the RNA-linked protein VpG). The formation of virus-antibody complexes of different stoichiometries (up to a mass 12.5 x 10(6) Da corresponding to 30 attached antibodies) and virus-receptor complexes (up to a mass 8.8 x 10(6) Da corresponding to 12 attached receptor molecules) was monitored. Via the volume derived from the electrophoretic mobility diameter (EMD), the stoichiometry of the HRV complexes was calculated. The accuracy of the EMD was within +/- 0.5 nm, which corresponds to an accuracy of +/- 4 antibodies and +/- 5 receptor molecules in the respective complexes. For the first time, we here demonstrate the use of nESI-GEMMA for the analysis of the size and stoichiometry of biomolecules in high-order complexes in real time under normal pressure conditions.