Unusually high mechanical stability of bacterial adhesin extender domains having calcium clamps

To gain insight into the relationship between protein structure and mechanical stability, single molecule force spectroscopy experiments on proteins with diverse structure and topology are needed. Here, we measured the mechanical stability of extender domains of two bacterial adhesins MpAFP and MhLap, in an atomic force microscope. We find that both proteins are remarkably stable to pulling forces between their N- and C- terminal ends. At a pulling speed of 1 μm/s, the MpAFP extender domain fails at an unfolding force F(u) = 348 ± 37 pN and MhLap at F(u) = 306 ± 51 pN in buffer with 10 mM Ca(2+). These forces place both extender domains well above the mechanical stability of many other β-sandwich domains in mechanostable proteins. We propose that the increased stability of MpAFP and MhLap is due to a combination of both hydrogen bonding between parallel terminal strands and intra-molecular coordination of calcium ions.