Kwon Seokmu, Andreas Michael P, Giessen Tobias W
Department of Chemical Engineering, University of Michigan, Ann Arbor, MI 48109, USA.
Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
bioRxiv. 2023 Jul 26:2023.07.26.550694. doi: 10.1101/2023.07.26.550694.
Encapsulins are self-assembling protein nanocompartments able to selectively encapsulate dedicated cargo enzymes. Encapsulins are widespread across bacterial and archaeal phyla and are involved in oxidative stress resistance, iron storage, and sulfur metabolism. Encapsulin shells exhibit icosahedral geometry and consist of 60, 180, or 240 identical protein subunits. Cargo encapsulation is mediated by the specific interaction of targeting peptides or domains, found in all cargo proteins, with the interior surface of the encapsulin shell during shell self-assembly. Here, we report the 2.53 Å cryo-EM structure of a heterologously produced and highly cargo-loaded T3 encapsulin shell from and explore the systems' structural heterogeneity. We find that exceedingly high cargo loading results in the formation of substantial amounts of distorted and aberrant shells, likely caused by a combination of unfavorable steric clashes of cargo proteins and shell conformational changes. Based on our cryo-EM structure, we determine and analyze the targeting peptide-shell binding mode. We find that both ionic and hydrophobic interactions mediate targeting peptide binding. Our results will guide future attempts at rationally engineering encapsulins for biomedical and biotechnological applications.
封装蛋白是能够选择性封装特定货物酶的自组装蛋白质纳米隔室。封装蛋白广泛存在于细菌和古菌门中，参与氧化应激抗性、铁储存和硫代谢。封装蛋白外壳呈现二十面体几何形状，由60、180或240个相同的蛋白质亚基组成。货物封装是由所有货物蛋白中发现的靶向肽或结构域在外壳自组装过程中与封装蛋白外壳内表面的特异性相互作用介导的。在这里，我们报告了一种来自异源产生且高负载货物的T3封装蛋白外壳的2.53 Å冷冻电镜结构，并探索了该系统的结构异质性。我们发现极高货物负载会导致大量扭曲和异常外壳的形成，这可能是由货物蛋白不利的空间冲突和外壳构象变化共同引起的。基于我们的冷冻电镜结构，我们确定并分析了靶向肽与外壳的结合模式。我们发现离子相互作用和疏水相互作用都介导靶向肽的结合。我们的结果将指导未来为生物医学和生物技术应用合理设计封装蛋白的尝试。
