Pore occupancy is modified by charge addition along the permeation pathway. (A) A snapshot of the Shaker pore structure and scheme of occupancy analysis. Shaker protein, shown in light pink, was modeled using the crystallographic structure of Kv1.2 channel as a template (PDB accession no. 2A79; Moscoso et al., 2012). Segments S5–S6, forming the pore domain, are the red ribbons; K+ are green spheres; lipids are in light gray in surf representation; and residues 471, 475, 476, and 479 are represented as gray, green, blue, and yellow bumps, respectively. The front and back subunits were removed for clarity. The discontinuous lines schematize the 5-Å-radius, pore-concentric cylinder in which time-averaged ion densities were calculated in 0.5-Å-thick slices. (B) Occupancy profile for Shaker-WT, V476D, S479D, A471D, P475D, P475D/V476D, P475D/S479D, and A471D/P475D/S479D calculated by the time-averaged ion densities inside the pore-concentric, 5-Å-radius cylinder relative to the bulk solution (500 mM K+) during ∼10 ns of simulation. Discontinuous lines indicate the binding sites at z of ∼14, ∼11, ∼8, ∼5, ∼2, approximately −9, approximately −15, and approximately −19 Å for s0, s1, s2, s3, s4, s5, s6, and s7, respectively. The drawing in A and B is at approximately the same Z-scale.
