Erwinia Pectic Enzyme Out System: A Model for Studying Exoprotein Secretion by Gram-Negative Bacteria

植物病原菌Erwinia chrysanhemi和E.carotovora的外源蛋白分泌系统(OUT)为研究蛋白质跨革兰氏阴性细菌两层膜的转运提供了一个有前途的实验模型。这种细菌选择性地分泌大量果胶酶和其他外源蛋白。一组菊花肠杆菌“out”基因使大肠杆菌能够分泌菊花肠杆菌外源蛋白。菊花杆菌的Out基因与肺炎克雷伯菌的普鲁兰酶分泌基因同源，这表明Erwinia模型将具有广泛的相关性。这两个欧文氏菌“OUT”系统可以方便地针对它们各自的外源蛋白。本项目的目的是通过DNA序列分析和Tn5tak1突变完成菊花肠杆菌“out”基因和编码蛋白的特征分析，鉴定控制外源蛋白分泌的胡萝卜肠杆菌和菊花肠杆菌“out”组分，并探索菊花肠杆菌与胡萝卜肠杆菌Pel1或β-内酰胺酶的杂交蛋白和缺失衍生物的分泌能力。对“Out”途径的充分理解可能导致干扰毒力蛋白的分泌或增强有益蛋白的分泌的一般技术。

The exoprotein secretion ("Out") system of the plant pathogens Erwinia chrysanthemi and E. carotovora provide a promising experimental model for studying the translocation of proteins across the two membranes that envelope gram-negative bacteria. The bacteria selectively secrete a remarkable array of pectic enzymes and other exoproteins. A cluster of E. chrysanthemi "out" genes enables Escherichia coli to secrete E. chrysanthemi exoproteins. The E. chrysanthemi "out" genes are homologous with the pullulanase secretion genes of Klebsiella pneumoniae, suggesting that the Erwinia model will be broadly relevant. The two Erwinia "out" systems are conveniently specific for their respective exoproteins. The objectives of this project are to complete the characterization of the E. chrysanthemi "out" genes and encoded proteins by DNA sequence analysis and Tn5tac1 mutagenesis, identify the E. carotovora and E. chrysanthemi "out" components controlling exoprotein secretion specificity, and explore the secretion ability of hybrid proteins and deletion derivatives involving E. chrysanthemi PelE and either E. carotovora Pel1 or beta-lactamase. An adequate understanding of the "out" pathway could lead to general techniques for either disrupting the secretion of virulence proteins or enhancing secretion of beneficial proteins.