Structural and functional characterization of TlyA hemolysin from Helicobacter pylori

Abstract

Helicobacter pylori gene product HP1086, designated as 'non-conventional' hemolysin named TlyA, is a virulence factor which plays an important role in persistent colonization and disease progression. In this study, 27-kDa TlyA from H. pylori was produced as a soluble recombinant protein in E. coli and purified to near homogeneity by cation exchange chromatography. Purified TlyA was biologically active as proven by membrane-perturbating activity against liposomes in vitro. Quaternary structural assembly and membrane-perturbing activity of TlyA negatively influenced by the addition of reducing agent DTT, thus suggesting a possible function of disulfide bonds for protein activity. Structural model suggests the potential formation of inter-molecular disulfide bridges between Cys124 and Cys128 residues located in the interface within the dimeric assembly. In addition, in silico modeling suggests S-adenosyl methionine (SAM) binding to TlyA could be inhibited by sinefungin which shares the similar binding residues with SAM. Recombinant TlyA causes cytotoxicity in human gastric cells and induces production of cytokine IL-8. Overall, this study disclosed an important role of intermolecular disulfide bonds structural assembly and membrane-perturbing activity of H. pylori TlyA.