DIPHTHERIA TOXIN AND ITS RECEPTOR

白喉毒素及其受体

• 批准号: 2060412
• 负责人: LEON EIDELS
• 金额: $ 24.19万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1980
• 资助国家: 美国
• 起止时间: 1980-04-01 至 1997-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2060412
• 关键词: Corynebacterium diphtheriae; antiidiotype antibody; antireceptor antibody; bacterial toxicology; diphtheria toxin; immunochemistry; laboratory mouse; laboratory rabbit; membrane activity; membrane proteins; microorganism immunology; molecular cloning; monoclonal antibody; nucleic acid sequence; protein structure; receptor; receptor binding; tissue /cell culture; transfection

The overall objective of this grant proposal is to characterize the receptor-binding domain- and the receptor-binding site within this domain - of diphtheria toxin, as well as to characterize fully the specific diphtheria toxin-binding cell surface protein(s) from highly toxin-sensitive Vero cells that function as the toxin receptor. The receptor-binding domain has recently been localized to the carboxyl- terminal Mr about 6,000 region of diphtheria toxin by our laboratory. The receptor-binding site will be localized further by chemical modifications o specific amino acid residues and by site- directed mutagenesis. Monoclonal antibodies to the toxin's receptor-binding domain/site will be prepared and will be employed to produce anti- idiotypic antibodies; the anti-idiotypic antibodies will then be tested for their ability to protect Vero Cells from diphtheria toxin-mediated cytotoxicity and for their ability to recognize the toxin receptor. Antibodies to the receptor will be employed to characterize and purify the receptor, and to investigate the cell surface distribution and mechanism of internalization of the toxin receptor on Vero cells. The diphtheria toxin-sensitive mouse cells, recently isolated by transfection of toxin-resistant mouse L-cells with DNA from Vero cells, will be employed to clone and sequence the toxin receptor; the sequence will be compared to that of other receptors, in order to determine the nature of the toxin receptor and/or its homology with other known receptors. The results of this proposed project will provide an insight into the possible nature of the physiological cell surface receptor that diphtheria toxin utilizes to gain illicit access to the cell cytosol, will significantly extend our knowledge on toxin:receptor interactions, and will further our understanding on receptor-mediated internalization of such macromolecules as growth factors, hormones, and other exotoxins.

该赠款提案的总体目的是表征 受体结合结构域和受体结合位点 域 - 白喉毒素的域，以及完全表征 来自高度的特异性白喉结合细胞表面蛋白 毒素敏感的VERO细胞起作用，可作为毒素受体。 最近已将受体结合域定位于羧基 - 我们的实验室大约6,000个白喉毒素的终端。 受体结合部位将通过化学物质进一步定位 修改o特定的氨基酸残基，并通过位置定向 诱变。毒素受体结合的单克隆抗体 将准备域/部位，并将用于生产抗 白痴型抗体；然后将测试抗IDiotypic抗体 因为它们能够保护Vero细胞免受白喉毒素介导 细胞毒性及其识别毒素受体的能力。 将使用对受体的抗体来表征和净化 受体，并研究细胞表面分布和 Vero细胞上毒素受体内在化的机理。这 白喉毒素敏感的小鼠细胞，最近通过 用Vero细胞的DNA转染抗毒素的小鼠L细胞， 将用来克隆并测序毒素受体；这 将序列与其他受体的序列进行比较，以便 确定毒素受体的性质和/或与其同源 其他已知的受体。 该提议的项目的结果将提供有关该项目的见解 生理细胞表面受体的可能性质 白喉毒素用来非法进入细胞细胞质， 将大大扩展我们对毒素的了解：受体相互作用， 并将进一步我们对受体介导的内在化的理解 在生长因子，激素和其他外毒素等大分子中。