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

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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.