Modification of the signal sequence cleavage site of listeriolysin O does not affect protein secretion but impairs the virulence of Listeria monocytogenes

Listeriolysin O (LLO, hly-encoded), a major virulence factor secreted by the bacterial pathogen Listeria monocytogenes, is synthesized as a precursor of 529 residues. To impair LLO secretion, the four residues of the predicted signal sequence cleavage site (EA-KD) were deleted and the mutant LLO protein was expressed in a hly-negative derivative of L. monocytogenes. Unexpectedly, the mutant protein was secreted in normal amounts in the culture supernatant and was fully haemolytic. N-terminal sequencing of the secreted LLO molecule revealed that N-terminal processing of the preprotein occurred three residues downstream of the natural cleavage site. L. monocytogenes expressing this truncated LLO showed a reduced capacity to disrupt the phagosomal membranes of bone marrow macrophages and of hepatocytes; and the mutant strain showed a 100-fold decrease in virulence in the mouse model. These results suggest that the first N-terminal residues of mature LLO participate directly in phagosomal escape and bacterial infection.

Molecular analysis of archaeal flagellins: similarity to the type IV pilin – transport superfamily widespread in bacteria

Ultrastructural, biochemical and genetic evidence has shown that the flagella and flagellin proteins from members of the archaea are distinct from their bacterial counterparts. The most important evidence is the sequence dissimilarity between archaeal and bacterial flagellins. We report here similarity between archaeal flagellins and members of the bacterial type IV pilin – transport superfamily. In addition to sequence similarity, the archaeal flagellins and the type IV pilin – transport superfamily share an unusual signal sequence cleavage site and may have functional parallels. This relationship has important implications for the assembly and biogenesis of archaeal flagella.Key words: flagellin, type IV pilin, homology, general secretion proteins, archaea.