BIOSYNTHESIS OF ENTEROBACTERIAL COMMON ANTIGEN

这项研究的长期目标是阐明这种机制。 参与黄瓜外膜成分的生物发生和功能 革兰氏阴性细菌。作为实现这一目标的途径，我们调查了 肠道细菌共同抗原(ECA)的生物化学和遗传学 在大肠杆菌和鼠伤寒沙门氏菌中合成。ECA是一个外在的 肠杆菌科特有的膜糖脂，存在于 属于这个家族的所有细菌。将特别强调 Eca生物合成机制的确定及鉴定和鉴定 描述参与这一过程的基因。这一努力一直是 促进大肠杆菌突变体的分离和鉴定 和鼠伤寒沙门氏菌在几个遗传决定因素中存在缺陷 非洲经委会。这些突变体的特征导致了 Eca合成和合成过程中生物合成中间体的鉴定 体外和体内实验系统的发展 Eca组装过程中涉及的特定酶步骤的演示。 为了继续我们的研究，我们在这里提议进行实验，并且 请求的支持期限的具体目标是：(I)确定 磷脂连接Eca的合成机制 多糖链(ECA-PL)，(Ii)确定参与的机制 Eca多糖链的组装，(Iii)确定其作用机制 参与将ECA多糖链转移到 脂多糖受体形成ECA-LPs，以及(Iv)研究 开放阅读框O348在Eca组装中的可能作用 分子。 这项拟议的研究将利用分子生物学技术， 生物化学和遗传学。预计，如果了解到 ECA组装的机制将提供对 细菌中膜的生物发生和组装。这样的信息将 为新型抗菌剂的开发提供了合理的依据， 它还将提供对参与 真核细胞和细胞膜的生物发生和组装 细胞器。

The long term goals of this research are to elucidate the mechanisms involved in the biogenesis and function of outer membrane components of gram-negative bacteria. As an approach to this goal, we have investigated the biochemistry and genetics of enterobacterial common antigen (ECA) synthesis in Escherichia coli and Salmonella typhimurium. ECA is an outer membrane glycolipid unique to the Enterobacteriaceae, and it is present in all bacteria belonging to this family. Specific emphasis will be placed on determining the mechanism of biosynthesis of ECA and on identifying and characterizing the genes involved in this process. This endeavor has been facilitated by the isolation and characterization of mutants of E. coli and S. typhimurium defective in several of the genetic determinants of ECA. The characterization of these mutants has resulted in the identification of biosynthetic intermediates involved in ECA synthesis and the development of in vitro and in vivo experimental systems that allow demonstration of specific enzymatic steps involved in ECA assembly. We propose here experiments for the continuation of our studies, and the specific aims for the requested period of support are to (i) determine the mechanism involved in the synthesis of phospholipid-linked ECA polysaccharide chains (ECA-PL), (ii) determine the mechanism involved in the assembly of ECA polysaccharide chains, (iii) determine the mechanism involved in the transfer of ECA polysaccharide chains to lipopolysaccharide acceptors to form ECA-LPs, and (iv) investigate the possible function of open reading frame o348 in the assembly of ECA molecules. The proposed research will utilize techniques of molecular biology, biochemistry, and genetics. It is anticipated that a knowledge of the mechanism of ECA assembly will provide insights into the processes of membrane biogenesis and assembly in bacteria. Such information will provide a rational basis for the development of new antimicrobial agents, and it will also provide insights into the mechanisms involved in the biogenesis and assembly of membranes in eukaryotic cells and cell organelles.