OPTIMIZING CELL SURFACE DISPLAY IN E COLI

优化大肠杆菌中的细胞表面显示

• 批准号: 2536424
• 负责人: WILLIAM JOSEPH COLEMAN
• 金额: $ 10万
• 依托单位: KAIROS SCIENTIFIC, INC.
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-09-30 至 1998-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2536424
• 关键词: Escherichia coli; bacterial proteins; cell membrane; chimeric proteins; flow cytometry; fluorimetry; green fluorescent proteins; membrane proteins; nonblood lipoprotein; polymerase chain reaction; protein engineering; protein sequence; protein signal sequence

DESCRIPTION: Surface display of foreign proteins on the outer membrane of bacterial cells provides a unique and potentially powerful means to screen for novel functionality. However, practical constraints on the implementation of this technology prevents its widespread application. One of the best characterized anchor proteins for display is a fusion between a fragment of the major E. coli lipoprotein (Lpp) and a fragment of the outer membrane protein, OmpA. Dr. Coleman proposes to use a fusion of Lpp-OmpA with the Green Fluorescent Protein (Lpp-OmpA-GFP) to help engineer a membrane anchor protein that is more stable in E. coli, has the optimum level of surface expression, allows multiple rounds of cell sorting, and which permits the bacteria to be grown to a higher cell density and at a higher temperature. An improved cell surface display system will allow a greater variety of proteins to be expressed under more manageable conditions, providing an impetus for the advancement of such diverse technologies as antibody development and enzyme evolution.

描述：外源蛋白在外膜上的表面展示 细菌细胞提供了一种独特且潜在强大的筛选手段 新颖的功能。然而，实际限制 该技术的实施阻碍了其广泛应用。 用于展示的最具有特征的锚定蛋白之一是融合蛋白 主要大肠杆菌脂蛋白 (Lpp) 片段和片段之间 外膜蛋白 OmpA。科尔曼博士建议使用 Lpp-OmpA 与绿色荧光蛋白 (Lpp-OmpA-GFP) 融合 帮助设计一种在大肠杆菌中更稳定的膜锚定蛋白， 具有最佳的表面表达水平，允许多轮 细胞分选，使细菌能够生长到更高的细胞 密度和更高的温度。改进的细胞表面显示 系统将允许更多种类的蛋白质在 更易于管理的条件，为进步提供动力 抗体开发和酶进化等多种技术。