Abstract
BackgroundOmptins are a family of outer membrane proteases that have spread by horizontal gene transfer in Gram-negative bacteria that infect vertebrates or plants. Despite structural similarity, the molecular functions of omptins differ in a manner that reflects the life style of their host bacteria. To simulate the molecular adaptation of omptins, we applied site-specific mutagenesis to make Epo of the plant pathogenic Erwinia pyrifoliae exhibit virulence-associated functions of its close homolog, the plasminogen activator Pla of Yersinia pestis. We addressed three virulence-associated functions exhibited by Pla, i.e., proteolytic activation of plasminogen, proteolytic degradation of serine protease inhibitors, and invasion into human cells.ResultsPla and Epo expressed in Escherichia coli are both functional endopeptidases and cleave human serine protease inhibitors, but Epo failed to activate plasminogen and to mediate invasion into a human endothelial-like cell line. Swapping of ten amino acid residues at two surface loops of Pla and Epo introduced plasminogen activation capacity in Epo and inactivated the function in Pla. We also compared the structure of Pla and the modeled structure of Epo to analyze the structural variations that could rationalize the different proteolytic activities. Epo-expressing bacteria managed to invade human cells only after all extramembranous residues that differ between Pla and Epo and the first transmembrane β-strand had been changed.ConclusionsWe describe molecular adaptation of a protease from an environmental setting towards a virulence factor detrimental for humans. Our results stress the evolvability of bacterial β-barrel surface structures and the environment as a source of progenitor virulence molecules of human pathogens.
Highlights
Omptins are a family of outer membrane proteases that have spread by horizontal gene transfer in Gram-negative bacteria that infect vertebrates or plants
Epo expressed by E. coli XL1 degraded plasminogen but significantly more slowly than did Pla expressed in the same host strain in a 2-h incubation with plasminogen (Figure 2B)
We have recently reported that Epo and Pla degrade the serpin plasminogen activator inhibitor 1 (PAI-1) [16], and recombinant E. coli with Pla or Epo cleaved another serpin, a2AP (Figure 2C)
Summary
Omptins are a family of outer membrane proteases that have spread by horizontal gene transfer in Gram-negative bacteria that infect vertebrates or plants. To simulate the molecular adaptation of omptins, we applied site-specific mutagenesis to make Epo of the plant pathogenic Erwinia pyrifoliae exhibit virulence-associated functions of its close homolog, the plasminogen activator Pla of Yersinia pestis. A conclusion from these studies is that highly similar genes, many of which are associated with bacterial virulence, are found across great phylogenetic distances and in different genetic elements, which is indicative of horizontal gene transfer. These families of virulence factors - including toxins, transport systems, adhesins, and antibiotic resistance factors - have evolved by adaptive radiation of a functional progenitor molecule to and within other strains and species to support survival in differing ecological. Pla is needed for establishment of bubonic plague [10,11], and a critical role of Pla has been described in pneumonic plague where it enables localized growth of Y. pestis in the lungs [12]
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