Subset of hybrid eukaryotic proteins is exported by the type I secretion system of Erwinia chrysanthemi

Erwinia chrysanthemi exports degradative enzymes by using a type I protein secretion system. The proteases secreted by this system lack an N-terminal signal peptide but contain a C-terminal secretion signal. To explore the su bstrate specificity of this system, we have expressed the E. chrysanthemi t ransporter system (prtDEF genes) in Escherichia coli and tested the ability of this ABC transporter to export hybrid proteins carrying C-terminal frag ments off. chrysanthemi protease B. The C terminus contains six glycine-ric h repeated motifs, followed by two repeats of the sequences DFLV and DW. Tw o types of hybrid proteins were assayed for transport, proteins with the 93 -residue-protease-B C terminus containing one glycine rich repeat and both hydrophobic terminal repeats and proteins with the 181-residue C terminus c ontaining all repeat motifs, Although the shorter C terminus is unable to e xport the hybrids, the longer C terminus can promote the secretion of hybri d proteins with N termini as large as 424 amino acids, showing that the gly cine-rich motifs are required for the efficient secretion of these hybrids. However, the secretion of hybrids occurs only if these proteins do not car ry disulfide bonds in their mature structures. These latter results suggest that disulfide bond formation can occur prior to or during the secretion. Disulfide bonds may prevent type I secretion of hybrids. One simple hypothe sis to explain these results is that the type I channel is too narrow to pe rmit the export of proteins with secondary structures stabilized by disulfi de bonds.