Fig. 3.

pH-dependent conversion of the T-domain from the soluble W-state into the membrane-competent W⁺-state. (A) Identification of W-to-W+ transition through membrane binding at lipid saturation. Mobility measurements based on FCS (diamonds) and measurements of FRET between donor-labeled T-domain and acceptor-labeled LUVs (circles) were performed as described in [Kyrychenko et al., 2009]. Triangles represent the increase in fluorescence intensity of environment-sensitive probe bimane attached to a single cysteine in position 369 (Fig. 2A). (B) Illustration of conformational changes associated with protonation of T-domain’s six histidines studied by MD simulations [Kurnikov et al., 2013]. The collapse of TH1–2 and exposure of TH8 are also confirmed by HDX-MS [Li, Rodnin, Ladokhin, & Gross, 2014; Rodnin et al., 2016]. Reprinted from Kyrychenko, A., Posokhov, Y.O., Rodnin, M.V., Ladokhin, A.S. (2009) Kinetic intermediate reveals staggered ph-dependent transitions along the membrane insertion pathway of the diphtheria toxin t-domain. Biochemistry, 48, 7584–7594; Copyright 2013, with permission from Elsevier.