PMC

Increased hydration in the extracellular half of the translocon upon displacement of the plug. The wild-type translocon (panel A; Sim1) is compared to the L406K mutant (panel B; Sim4). Color code used for van der Waals spheres: sidechains of the amino acids forming the hydrophobic pore - mauve, except for L406 which is depicted in violet; sidechains of S65 and A64 - green; water oxygen atoms – yellow; lipid nitrogen, oxygen, phosphorous, and carbon atoms are depicted in blue, red, orange, and black, respectively. For simplicity, hydrogen atoms are not shown. (C) Close-up view of the hydrophobic pore in the wild-type (Sim1; green) and L406K (orange; Sim4). Sidechains of pore ring amino acids and A64 are shown as van der Waals spheres; water molecules within 6 Å of A64 and within 6 Å of L/K406 are shown as cyan and purple spheres, respectively. (D) Histograms of the number of water molecules within 6 Å of A64 in the wild-type (Sim1; green), K250E (Sim2; purple), L406K (orange, Sim4), and E336R (Sim5). The histograms were computed with a bin size of 1.

Determination of ligand-arrestin-1 dissociation constants from the NMR titration data. (A) Chemical shift changes of residues (A64, I16, L172, Y67) are plotted as a function of IP6 concentration, to which was fit a 1:1 binding model using NMRView titration analysis module to obtain dissociation constants. (B) Chemical shift changes of residues (A64, I16, L172, Y67) are plotted as a function of heparin concentration, with the same analysis of the data and the determination of apparent dissociation constants K_d,app being performed as in (A).

Schematic of intragenic sequence rearrangements for A64 genes 7, 10, and 11. Each aberrant RNA segment contains a single head-to-tail sequence duplication (DUP-1 or DUP-2). The duplication initiates downstream of the ORF for each gene, thereby allowing A64 to express full-length NSP3, NSP4, and NSP5/6 proteins, respectively. The gene 11 rearrangement introduces a second NSP6 ORF, one that is positioned downstream of the NSP5 ORF.

PFGE filters probed with MPHx. The two filters (lanes 1 to 6 and 7 to 11, respectively) were probed with MPHx and then exposed for 3 days. The samples were as follows: lane 1, S150-2B; lane 2, CEN.PK2-1D; lane 3, RH144-3A; lane 4, A15; lane 5, A64; lane 6,A60; lane 7, A24; lane 8, A64; lane 9, A72; lane 10, A179; and lane 11, A180. The positions of chromosomes IV and X, the duplex VII/XV, and the sample slot are shown.