<br>类弹性蛋白多肽 (ELP) 电荷影响 ELP-mCherry 融合蛋白的自组装,Biomacromolecules

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类弹性蛋白多肽 (ELP) 电荷影响 ELP-mCherry 融合蛋白的自组装
Biomacromolecules ( IF 5.5 ) Pub Date : 2018-05-23 00:00:00 , DOI: 10.1021/acs.biomac.8b00147
Carolyn E Mills ₁ , Zachary Michaud ₁ , Bradley D Olsen ₁

Affiliation

蛋白质-聚合物生物共轭物的自组装提供了一种控制功能材料中球状蛋白质的纳米结构和方向的优雅策略。最近的研究表明，球状蛋白 mCherry 与弹性蛋白样多肽 (ELP) 的基因融合产生了与这些蛋白质-聚合物生物缀合物相似的自组装行为。在研究蛋白质-聚合物生物共轭物自组装的背景下，ELP 序列的可变性允许正交调整 ELP 块的几种不同特性，同时保持一致的多肽主链化学。这项工作利用这种 ELP 序列可调性与氨基酸序列基因工程提供的精确控制相结合，生成了四个新型 ELP 序列的文库，用于研究电荷和疏水性对 ELP-mCherry 融合蛋白自身的综合影响-集会。通过小角 X 射线散射 (SAXS) 和去偏光散射 (DPLS) 研究浓溶液自组装。这些实验表明，含有带负电荷的 ELP 块的融合体根本不会组装，而带有电荷平衡的 ELP 块的融合体表现出较弱的组装倾向。相比之下，含有不带电 ELP 块的融合蛋白在溶液中浓度为 40 wt% 时开始有序化，并且在所有测量浓度下都具有比其他融合蛋白更尖锐、更强烈的 SAXS 峰。这些实验表明，ELP 嵌段的电荷特性比 ELP 嵌段的疏水性更能预测自组装行为。仅 ELP 上的稀溶液小角中子散射 (SANS) 表明本研究中使用的所有 ELP（包括不带电的 ELP）均采用与传统聚合物（包括聚两性电解质和聚电解质）相似的稀溶液构象。最后，对 ELP-mCherry 混合物的动态光散射研究表明，带电 ELP 和 mCherry 之间不存在明显的络合作用。因此，有人提出，含有不带电 ELP 块的融合蛋白的优异自组装归因于带电和不带电块之间由于局部电荷相关效应而产生的有效排斥，并且在阴离子 ELP 的情况下，ELP 内相同电荷之间的排斥堵塞。

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Elastin-like Polypeptide (ELP) Charge Influences Self-Assembly of ELP–mCherry Fusion Proteins

Self-assembly of protein–polymer bioconjugates presents an elegant strategy for controlling nanostructure and orientation of globular proteins in functional materials. Recent work has shown that genetic fusion of globular protein mCherry to an elastin-like polypeptide (ELP) yields similar self-assembly behavior to these protein–polymer bioconjugates. In the context of studying protein–polymer bioconjugate self-assembly, the mutability of the ELP sequence allows several different properties of the ELP block to be tuned orthogonally while maintaining consistent polypeptide backbone chemistry. This work uses this ELP sequence tunability in combination with the precise control offered by genetic engineering of an amino acid sequence to generate a library of four novel ELP sequences that are used to study the combined effect of charge and hydrophobicity on ELP–mCherry fusion protein self-assembly. Concentrated solution self-assembly is studied by small-angle X-ray scattering (SAXS) and depolarized light scattering (DPLS). These experiments show that fusions containing a negatively charged ELP block do not assemble at all, and fusions with a charge balanced ELP block exhibit a weak propensity for assembly. By comparison, the fusion containing an uncharged ELP block starts to order at 40 wt % in solution and at all concentrations measured has sharper, more intense SAXS peaks than other fusion proteins. These experiments show that charge character of the ELP block is a stronger predictor of self-assembly behavior than the hydrophobicity of the ELP block. Dilute solution small-angle neutron scattering (SANS) on the ELPs alone suggests that all ELPs used in this study (including the uncharged ELP) adopt dilute solution conformations similar to those of traditional polymers, including polyampholytes and polyelectrolytes. Finally, dynamic light scattering studies on ELP–mCherry blends shows that there is no significant complexation between the charged ELPs and mCherry. Therefore, it is proposed that the superior self-assembly of fusion proteins containing uncharged ELP block is due to effective repulsions between charged and uncharged blocks due to local charge correlation effects and, in the case of anionic ELPs, repulsion between like charges within the ELP block.

更新日期：2018-05-23

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