Department of Molecular Genetics and Microbiology, University of New Mexico School of Medicine, Albuquerque, New Mexico 87131, USA.
J Nanobiotechnology. 2011 May 24;9:22. doi: 10.1186/1477-3155-9-22.
To be useful for genetic display of foreign peptides a viral coat protein must tolerate peptide insertions without major disruption of subunit folding and capsid assembly. The folding of the coat protein of RNA phage MS2 does not normally tolerate insertions in its AB-loop, but an engineered single-chain dimer readily accepts them as long as they are restricted to one of its two halves.
Here we characterize the effects of peptide insertions on the thermal stabilities of MS2 virus-like particles (VLPs) displaying a variety of different peptides in one AB-loop of the coat protein single-chain dimer. These particles typically denature at temperatures around 5-10°C lower than unmodified VLPs. Even so, they are generally stable up to about 50°C. VLPs of the related RNA phage PP7 are cross-linked with intersubunit disulfide bonds and are therefore significantly more stable. An AB-loop insertion also reduces the stability of PP7 VLPs, but they only begin to denature above about 70°C.
VLPs assembled from MS2 single-chain dimer coat proteins with peptide insertions in one of their AB-loops are somewhat less stable than the wild-type particle, but still resist heating up to about 50°C. Because they possess disulfide cross-links, PP7-derived VLPs provide an alternate platform with even higher stability.
为了对外源肽进行遗传展示，病毒外壳蛋白必须能够容忍肽插入，而不会严重破坏亚基折叠和衣壳组装。RNA 噬菌体 MS2 的外壳蛋白的折叠通常不能容忍其 AB 环中的插入，但工程化的单链二聚体只要限制在其两个半体之一中，就可以很容易地接受它们。
在这里，我们研究了在 MS2 病毒样颗粒 (VLPs) 中展示各种不同肽的外壳蛋白单链二聚体的 AB 环中的肽插入对其热稳定性的影响。这些颗粒通常在未修饰的 VLPs 变性温度低约 5-10°C 的温度下变性。即便如此，它们通常在约 50°C 以下是稳定的。相关的 RNA 噬菌体 PP7 的 VLPs 与亚基间二硫键交联，因此稳定性显著提高。AB 环插入也会降低 PP7 VLPs 的稳定性，但它们仅在约 70°C 以上才开始变性。
用单链二聚体外壳蛋白组装的 MS2 病毒样颗粒，其 AB 环中的一个环插入了肽，其稳定性比野生型颗粒略低，但仍能抵抗加热至约 50°C。由于它们具有二硫键交联，因此源自 PP7 的 VLPs 提供了一个具有更高稳定性的替代平台。
