Two-Dimensional Crystallization of P22 Virus-Like Particles

详细信息    查看全文

• 作者：Hideyuki Yoshimura ; Ethan Edwards ; Masaki Uchida ; Kimberly McCoy ; Raj Roychoudhury ; Benjamin Schwarz ; Dustin Patterson ; Trevor Douglas
• 刊名：Journal of Physical Chemistry B
• 出版年：2016
• 出版时间：July 7, 2016
• 年：2016
• 卷：120
• 期：26
• 页码：5938-5944
• 全文大小：412K
• 年卷期：0
• ISSN：1520-5207

文摘

Virus-like particles (VLPs) are well established platforms for constructing functional biomimetic materials. The VLP from the bacteriophage P22 can be used as a nanocontainer to sequester active enzymes, at high concentration, within its cavity through a process of directed self-assembly. Construction of ordered 2D assemblies of these catalytic VLPs can be envisioned as a functional membrane. To achieve this, it is important to establish methods to fabricate densely packed monolayers of VLPs. Highly ordered assemblies of P22 can also be utilized as a two-dimensional (2D) crystal for electron crystallography to get precise structural information on the VLP. Here we report 2D crystallization of different P22 morphologies: P22 procapsid (PC), enzyme encapsulated PC (β-glycosidase and enhanced green fluorescent protein), empty shell (PC without scaffold proteins, ES), the expanded form of P22 (EX), and enzyme encapsulated EX (NADH oxidase). The 2D crystals of P22 VLPs were formed on a positively charged lipid monolayer at the water–air interface with a subphase containing 1% trehalose. A P22 solution, injected underneath the lipid monolayer, floated to the surface because of the density difference between the subphase and protein solution. The lipid monolayer, with adsorbed P22, was transferred to a holey carbon grid and was examined by electron microscopy. 2D crystals were obtained from a subphase containing 100 mM NaCl, 10 mM MES (pH 5.0), and 1% trehalose. The diffraction spots from the transferred film extended to the sixth order in negatively stained samples and the 10th order in cryo-electron microscopy samples.