Fig. 3: Molecular dynamics simulations-guided mutational analysis of MHAS2168.

a The coarse-grained molecular dynamics simulation system of MHAS2168 in the outward-facing conformation. The mycobacterial plasma membrane lipid components are shown on the right, with headgroups colored blue. Solvent is not shown for clarity. b Density profile of TAGs along the membrane normal, centered with respect to the membrane midplane. The blue dotted lines are the average position of the phosphate headgroups. c The average number of TAG contacts from the MHAS2168^OUT (left) and MHAS2168^IN (right) simulations, respectively, projected on the surface of the protein and colored from blue (no contacts) to red (large number of contacts). d Vancomycin sensitivity assays in M. smegmatis dKO cells, complemented with wild type LprG/Rv1410 operon (WT_Mtb), or mutant operons containing unaltered LprG (Rv1411) and mutated transporter Rv1410 as indicated. e Analogous analysis as in (d), with M. smegmatis dKO cells expressing instead wild type MHAS2167/68 operon (WT_MH) or mutant operons containing unaltered LprG (MHAS2167) and mutated transporter MHAS2168 as indicated. Δβ-HP – β-hairpin truncation; ΔAB - truncation of linker helices TMA and TMB. The growth curves in (d) and (e) are representative of three biological replicates and data are presented as mean values +/- SD of four technical replicates.