1887

Abstract

The last ORF of an gene cluster, designated , is required for the secretion of extracellular enzymes across the outer membrane in pv. It could encode a protein of 759 amino acid residues. A consensus N-terminal lipoprotein signal peptide was revealed from its deduced amino acid sequence. A [H]palmitate labelling experiment indicated that XpsD was fatty-acylated. Differential extraction with Triton X-100 disclosed that XpsD was fractionated with the outer membrane. Sucrose gradient sedimentation analysis of total membranes also indicated that XpsD was mainly located in the outer membrane. At least part of XpsD is exposed to the cell surface as suggested by trypsin experiment results. Intact cells pretreated with antibody against XpsD could indirectly be labelled with fluorescent agent. When the N-terminal lipoprotein signal peptide was replaced with a nonlipoprotein signal peptide cleavable by signal peptidase I, non-fatty-acylated XpsD was synthesized. Its subcellular location was indistinguishable from that of the fatty-acylated XpsD. Complementation of an ::Tn5 mutant of pv. indicated that this non-fatty-acylated XpsD remains functional in extracellular protein secretion. A stable, C-terminal truncated protein, XpsDd414-759, was synthesized from a mutated gene. Although it stayed associated with the outer membrane and exposed to the cell surface, it no longer could complement the ::Tn5 mutant of pv.

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1995-06-01
2021-10-17
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