Biomimetic Silica Encapsulation of Lipid Nanodiscs and β-Sheet-Stabilized Diacylglycerol Kinase,Bioconjugate Chemistry

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Biomimetic Silica Encapsulation of Lipid Nanodiscs and β-Sheet-Stabilized Diacylglycerol Kinase
Bioconjugate Chemistry ( IF 4.0 ) Pub Date : 2021-07-21 , DOI: 10.1021/acs.bioconjchem.1c00260
Friedrich Bialas ₁ , Christian F W Becker ₁

Affiliation

Integral membrane proteins (IMPs) comprise highly important classes of proteins such as transporters, sensors, and channels, but their investigation and biotechnological application are complicated by the difficulty to stabilize them in solution. We set out to develop a biomimetic procedure to encapsulate functional integral membrane proteins in silica to facilitate their handling under otherwise detrimental conditions and thereby extend their applicability. To this end, we designed and expressed new fusion constructs of the membrane scaffold protein MSP with silica-precipitating peptides based on the R5 sequence from the diatom Cylindrotheca fusiformis. Transmission electron microscopy (TEM) and atomic force microscopy (AFM) revealed that membrane lipid nanodiscs surrounded by our MSP variants fused to an R5 peptide, so-called nanodiscs, were formed. Exposing them to silicic acid led to silica-encapsulated nanodiscs, a new material for stabilizing membrane structures and a first step toward incorporating membrane proteins in such structures. In an alternative approach, four fusion constructs based on the amphiphilic β-sheet peptide BP-1 and the R5 peptide were generated and successfully employed toward silica encapsulation of functional diacylglycerol kinase (DGK). Silica-encapsulated DGK was significantly more stable against protease exposure and incubation with simulated gastric fluid (SGF) and intestinal fluid (SIF).

中文翻译：

脂质纳米盘和 β-片层稳定的二酰基甘油激酶的仿生二氧化硅封装

整合膜蛋白 (IMP) 包含非常重要的一类蛋白质，例如转运蛋白、传感器和通道，但由于难以在溶液中稳定它们，因此它们的研究和生物技术应用变得复杂。我们着手开发一种仿生程序，将功能性整合膜蛋白封装在二氧化硅中，以促进它们在其他有害条件下的处理，从而扩展它们的适用性。为此，我们设计并表达了膜支架蛋白 MSP 与基于硅藻Cylindrotheca fusiformis的 R5 序列的二氧化硅沉淀肽的新融合构建体。透射电子显微镜 (TEM) 和原子力显微镜 (AFM) 显示，形成了由与 R5 肽融合的 MSP 变体包围的膜脂质纳米盘，即所谓的纳米盘。将它们暴露于硅酸中产生了二氧化硅封装的纳米圆盘，这是一种用于稳定膜结构的新材料，也是将膜蛋白纳入此类结构的第一步。在另一种方法中，生成了四种基于两亲性 β-折叠肽 BP-1 和 R5 肽的融合构建体，并成功用于功能性二酰基甘油激酶 (DGK) 的二氧化硅封装。二氧化硅封装的 DGK 对于蛋白酶暴露以及与模拟胃液 (SGF) 和肠液 (SIF) 的孵育显着更稳定。

更新日期：2021-08-19

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