Mechanical forces regulate the reactivity of a thioester bond in a bacterial adhesin

• Published in: The Journal of biological chemistry Vol. 292; no. 21; pp. 8988 - 8997
• Main Authors: Echelman, Daniel J., Lee, Alex Q., Fernández, Julio M.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 26.05.2017; American Society for Biochemistry and Molecular Biology
• Subjects: adhesin; Adhesins, Bacterial - chemistry; atomic force microscopy (AFM); bacterial adhesion; Disulfides - chemistry; Editors' Picks; Gram-positive bacteria; Histamine - chemistry; Methylamines - chemistry; pilus mechanics; Protein Folding; single-molecule biophysics; Streptococcus pyogenes - chemistry; thioester bond; single-molecule biophysics; adhesin; atomic force microscopy (AFM); thioester bond; Gram-positive bacteria; pilus mechanics; bacterial adhesion
• ISSN: 0021-9258; 1083-351X; 1083-351X
• DOI: 10.1074/jbc.M117.777466

Bacteria must withstand large mechanical shear forces when adhering to and colonizing hosts. Recent structural studies on a class of Gram-positive bacterial adhesins have revealed an intramolecular Cys-Gln thioester bond that can react with surface-associated ligands to covalently anchor to host surfaces. Two other examples of such internal thioester bonds occur in certain anti-proteases and in the immune complement system, both of which react with the ligand only after the thioester bond is exposed by a proteolytic cleavage. We hypothesized that mechanical forces in bacterial adhesion could regulate thioester reactivity to ligand analogously to such proteolytic gating. Studying the pilus tip adhesin Spy0125 of Streptococcus pyogenes, we developed a single molecule assay to unambiguously resolve the state of the thioester bond. We found that when Spy0125 was in a folded state, its thioester bond could be cleaved with the small-molecule nucleophiles methylamine and histamine, but when Spy0125 was mechanically unfolded and subjected to forces of 50–350 piconewtons, thioester cleavage was no longer observed. For folded Spy0125 without mechanical force exposure, thioester cleavage was in equilibrium with spontaneous thioester reformation, which occurred with a half-life of several minutes. Functionally, this equilibrium reactivity allows thioester-containing adhesins to sample potential substrates without irreversible cleavage and inactivation. We propose that such reversible thioester reactivity would circumvent potential soluble inhibitors, such as histamine released at sites of inflammation, and allow the bacterial adhesin to selectively associate with surface-bound ligands.