Castellanos Milagros, Carrillo Pablo J P, Mateu Mauricio G
Centro de Biología Molecular "Severo Ochoa" (CSIC-UAM), Universidad Autónoma de Madrid, Madrid 28049, Spain.
Nanoscale. 2015 Mar 19;7(13):5654-64. doi: 10.1039/c4nr07046a.
Viruses are increasingly being studied from the perspective of fundamental physics at the nanoscale as biologically evolved nanodevices with many technological applications. In viral particles of the minute virus of mice (MVM), folded segments of the single-stranded DNA genome are bound to the capsid inner wall and act as molecular buttresses that increase locally the mechanical stiffness of the particle. We have explored whether a quantitative linkage exists in MVM particles between their DNA-mediated stiffening and impairment of a heat-induced, virus-inactivating structural change. A series of structurally modified virus particles with disrupted capsid-DNA interactions and/or distorted capsid cavities close to the DNA-binding sites were engineered and characterized, both in classic kinetics assays and by single-molecule mechanical analysis using atomic force microscopy. The rate constant of the virus inactivation reaction was found to decrease exponentially with the increase in elastic constant (stiffness) of the regions closer to DNA-binding sites. The application of transition state theory suggests that the height of the free energy barrier of the virus-inactivating structural transition increases linearly with local mechanical stiffness. From a virological perspective, the results indicate that infectious MVM particles may have acquired the biological advantage of increased survival under thermal stress by evolving architectural elements that rigidify the particle and impair non-productive structural changes. From a nanotechnological perspective, this study provides proof of principle that determination of mechanical stiffness and its manipulation by protein engineering may be applied for quantitatively probing and tuning the conformational dynamics of virus-based and other protein-based nanoassemblies.
作为具有多种技术应用的生物进化纳米器件，病毒越来越多地从纳米尺度的基础物理学角度进行研究。在小鼠微小病毒（MVM）的病毒颗粒中，单链DNA基因组的折叠片段与衣壳内壁结合，并作为分子支撑物，局部增加颗粒的机械刚度。我们探究了在MVM颗粒中，其DNA介导的硬化与热诱导的病毒失活结构变化的损伤之间是否存在定量联系。通过经典动力学分析以及使用原子力显微镜的单分子力学分析，对一系列衣壳 - DNA相互作用被破坏和/或靠近DNA结合位点的衣壳腔变形的结构修饰病毒颗粒进行了设计和表征。发现病毒失活反应的速率常数随着靠近DNA结合位点区域的弹性常数（刚度）增加而呈指数下降。过渡态理论的应用表明，病毒失活结构转变的自由能垒高度随局部机械刚度线性增加。从病毒学角度来看，结果表明感染性MVM颗粒可能通过进化使颗粒变硬并损害非生产性结构变化的结构元件，获得了在热应激下提高存活率的生物学优势。从纳米技术角度来看，这项研究提供了原理证明，即通过蛋白质工程确定机械刚度及其操纵可用于定量探测和调节基于病毒和其他基于蛋白质的纳米组装体的构象动力学。
