D-Protein and D-Peptide Technology in Applied Research

D-蛋白质和D-肽技术在应用研究中的应用

• 批准号: 2107414
• 金额: --
• 依托单位: CARDIFF UNIVERSITY
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2018
• 资助国家: 英国
• 起止时间: 2018 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2107414
• 关键词: Protein; Peptide; Technology; Applied; Research

Chemical protein synthesis provides access to entire D-protein enantiomers, with unique utility in applied research. Firstly, proteins may be crystallized from a racemic mixture of L- and D-enantiomers, known as racemic protein crystallography, providing facile access to high resolution X-ray structures. In this thesis, to understand how membrane-active bacteriocins have acquired their antibacterial properties, an analysis based on racemic protein crystallography (0.9-1.2 Å) of two key representatives, aureocin a53 (AucA) and lacticin Q (LnqQ) was conducted. Through structural analysis and systematic residue substitutions, conserved surface tryptophans appeared to play important roles in coordination of lipid phosphate and were critical for antibacterial activity. Additional tryptophan's in AucA were also involved in forming an oligomeric assembly for stability enhancement. Collectively, racemic protein crystallography shed light on the molecular interactions of tryptophans, demonstrating how a bacteriocin delivers its antibacterial properties. Secondly, D-enantiomers of protein drug targets can be used in mirror-image phage display (MIPD), allowing discovery of non-proteolytic D-peptide ligands as lead candidates. Development of a D-peptide capable of targeting tumor necrosis factor receptor I (TNFR1) activation could be a beneficial solution in controlling inflammation caused by cytokine storm. However, containing 144 residues and 12 disulfide bonds, synthesis of the D-enantiomer for MIPD would be a significant challenge. Here, isolation of the mid-chain, cytokine binding domain (TNRCD2) is reported, and its enantiomer enabled MIPD to discover a cyclic peptide binder with low micromolar affinity. Additionally, the use of computational methods to develop D-protein ligands based only on the target structure was explored.

化学蛋白质合成提供了获得整个d蛋白对映体的途径，在应用研究中具有独特的用途。首先，蛋白质可以从L-和d -对映体的外消旋混合物中结晶，称为外消旋蛋白晶体学，为高分辨率x射线结构提供了方便。为了了解膜活性细菌素是如何获得其抗菌特性的，本文基于消旋蛋白晶体学（0.9-1.2 Å）对两个关键代表，aureocin a53 （AucA）和lacticin Q （LnqQ）进行了分析。通过结构分析和系统残基取代，保守的表面色氨酸在脂质磷酸的协调中发挥重要作用，对抗菌活性至关重要。AucA中额外的色氨酸也参与形成寡聚体以增强稳定性。总的来说，外消旋蛋白晶体学揭示了色氨酸的分子相互作用，展示了细菌素如何传递其抗菌特性。其次，蛋白质药物靶点的d -对映体可用于镜像噬菌体展示（MIPD），从而发现非蛋白水解的d肽配体作为主要候选体。能够靶向肿瘤坏死因子受体I （TNFR1）激活的d肽的开发可能是控制细胞因子风暴引起的炎症的有益解决方案。然而，由于含有144个残基和12个二硫键，合成用于MIPD的d -对映体将是一个重大挑战。本文报道了中链细胞因子结合域（TNRCD2）的分离，其对映体使MIPD发现了具有低微摩尔亲和力的环肽结合物。此外，利用计算方法开发仅基于目标结构的d蛋白配体进行了探索。