Fig. 1.

Staphylococcal B domains are extremely resilient to mechanical force. a B domains, here from SdrG, are the link between the extremely strong interaction between the tip adhesin domain N2N3 (blues) binding a peptide (orange) presented by a host protein covering a surface (in this case fibrinogen, purple). b SdrG gene (top) and schematic (bottom): N-terminal N1 domain may be cleaved proteolytically, followed by the N2 and N3 domain that bind Fgß, B1 (green), and B2 (brown) each coordinate three Ca²⁺ ions and connect to the SD repeat region that gives the adhesin its name. The adhesin is covalently anchored to the bacterium’s peptidoglycan via a sortase motif (red). c MD equilibrated structure of the SdrG N2N3 domains connected to the fully ß-sheet Ig-like folds of the B1 and B2 domain (modeled from the homolog SdrD B1), each B domain coordinates three Ca²⁺ ions (yellow). d AFM-SMFS assay: covalent surface anchoring through polyethylene glycol (PEG) via the ybbr-tag (purple) using Fgß (orange)-ddFLN4 (cyan)-ybbr to probe SdrG N2N3-B1-ybbr on the cantilever. e Single force-extension trace at 0.8 µm s⁻¹ with unfolding of the ddFLN4 fingerprint (cyan arrow) at ~ 100 pN, followed by the SdrG B1 domain (green circle) at >2 nN (with the expected contour length increment of ~ 36 nm). Finally the SdrG N2N3:Fgß complex dissociates, allowing the cantilever to relax to zero force. f Dynamic force spectrum for the unfolding of the SdrG B1 domain at retraction velocities: 0.4 µm s⁻¹ (triangles, N = 574), 0.8 µm s⁻¹ (squares, N = 742), 1.6 µm s⁻¹ (diamonds, N = 878), 3.2 µm s⁻¹ (forward triangles, N = 789), 6.4 µm s⁻¹ (circles, N = 729). The high N value suggests that the B1 domain refolds on the cantilever. A Bell–Evans model fit (dashed line, ∆x = 0.082 nm, k_off⁰ = 3.8E–17 s⁻¹) through the most probable rupture force and force loading rate per velocity (large open markers, errors given as full-width at half maximum for each distribution) confirms the expected log-linear behavior