Fig. 4: Positively-charged surface residues on MlaC are responsible for mediating MlaA interaction.

a Cartoon and surface vacuum electrostatics representations of apo-MlaC (PDB 6GKI)^25. revealing positively-charged patches (blue surface). Positively-charged residues (or groups of residues, blue sticks) are numbered from 0 to 7. b Representative immunoblots showing abolishment of disulfide crosslinks between MlaA and MlaC in ΔmlaA ΔmlaC cells expressing indicated surface charged-reversed mutants (numbered according to (a)) of cysteine-substituted MlaC-His and MlaA-His from pET22/42 and pCDF plasmids, respectively. The experiment had been performed at least three times with similar results. Source data are provided as a Source Data file. c Representative immunoblots showing restoration of disulfide crosslinks between MlaA and MlaC in ΔmlaA ΔmlaC cells expressing charge-reversed MlaC1-His and MlaA^2DD2R-His from pET22/42 and pCDF plasmids, respectively. Samples were subjected to non-reducing (top) or reducing (bottom) SDS-PAGE prior to immunoblotting. This experiment had been performed at least three times with similar results. Source data are provided as a Source Data file. d Cartoon representations of Alphafold2-predicted MlaA (teal)-MlaC (pink) model, revealing the proximity of residues D243/D244 on MlaA, and residues K84/R90 on MlaC illustrated in blue and red sticks on MlaA and MlaC respectively (inset). Cutaway surface representation of this MlaA-MlaC model is shown on the right, revealing the juxtaposition of the hydrophobic lipid-binding cavity of MlaC with the hydrophilic channel of MlaA.