VIRUS BASED BIO-IMAGING & THERAPEUTIC DELIVERY SYSTEMS

许多病毒形成稳定的自组装蛋白质笼，其功能是储存、保护和运输核酸。 我们之前已经表明，病毒的笼状结构可以用作约束反应容器，用于封装和释放除其天然RNA基因组以外的各种材料。通过这种方式，病毒蛋白笼可以被认为是纳米尺寸的容器，能够通过明确的化学相互作用包裹其他分子。 目前的建议将探索使用这些病毒笼结构的封装和靶向输送的治疗剂，以及这些笼子的发展作为磁共振成像造影剂的基础上，我们证明的能力，工程的外壳蛋白。本提案的主要目的是开发一种模型病毒系统，用于在治疗材料（分子和聚合物）的高密度包装和释放中使用病毒笼结构。 病毒内的包装可以由内部蛋白质界面和相关治疗材料之间的静电互补驱动。 一个目的是通过工程化蛋白质笼内表面的静电性质来扩展可被截留在病毒蛋白质笼内的治疗材料的范围。第二个主要目的是开发病毒蛋白笼作为潜在的磁共振成像造影剂，通过工程化的病毒粒子上的固有金属结合位点结合180分子的顺磁性Gd（III）离子。第三个主要目的是在病毒体的外表面上表达来自层粘连蛋白的肽11，并确定其在特异性靶向表达层粘连蛋白结合蛋白的细胞的病毒笼中的有效性。第四个主要目的是利用病毒体中固有的结构转变来设计新的明确定义的化学开关（基于氧化还原电位和pH），以诱导用于选择性截留和释放治疗材料的门控。病毒体门控导致蛋白质笼中60个单独的20埃单位孔的可逆打开/关闭。 我们建议使用定点诱变工程二硫键和改变pH门控开关，在这些孔和测试他们的能力，捕获和释放治疗材料。

Many viruses form stable self-assembled protein cages that function to store, protect and transport nucleic acid. We have previously shown that the cage structure of viruses can be used as constrained reaction vessels for the encapsulation and release of a wide range of materials other than its native RNA genome. In this way virus protein cages can be thought of as nanometer sized containers able to encapsulate other molecules through well-defined chemical interactions. The current proposal will explore the use of these virus cage structures for encapsulation and targeted delivery of therapeutic agents as well as development of these cages as magnetic resonance imaging contrast agents based on our demonstrated ability to engineer the coat protein. The principle objective of this proposal is to develop a model viral system for the use of virus cage structures in the high- density packing and release of therapeutic materials (molecules and polymers). Packaging within the virus can be driven by electrostatic complementarily between the inner protein interface and the relevant therapeutic material(s). One objective will be to extend the range of therapeutic materials that can be entrapped within the viral protein cage by engineering the electrostatic properties of the inner surface of the protein cage. A second major objective is to develop viral protein cages as potential magnetic resonance imaging contrast agents by engineering the inherent metal binding sites on the virion for binding 180 molecules of the paramagnetic Gd(III) ion. A third major objective is to express peptide 11 from the laminin protein on the outer surface of the virion and to determine its effectiveness at specifically targeting viral cages to cells expressing laminin-binding protein. A fourth major objective is to utilize inherent structural transitions in the virion to engineer new well defined chemical switches (based on redox potential and pH) to induce gating for selective entrapment and release of therapeutic materials. Virion gating results in the reversible opening/closing of 60 separate 20Angstrom units holes in the protein cage. We propose to use site-directed mutagenesis to engineer disulfide linkages and altered pH gating switches at these pores and test for their ability to entrap and release therapeutic materials.