Fig. 2

mDAP and iGln interacting residues influence the survival. a Model of Mtb PknB PASTA3-4 dimer in complex with the muropeptide ligand. Inset panel shows a magnified view of the interactions between the mDAP and iGln residues of the muropeptide with polar amino acids between the PASTA3 and 4 domains of PknB. b Schematic representation of extra-cytoplasmic PASTA domain of PknB and the residues in the PASTA3-4 linker region suggested to be involved in interaction with iGln (SK) and mDAP (NQ) residues in the muropeptide. The mutations introduced in the linker region to generate PknB-GM are indicated. c Western blot analysis of WCLs prepared from Rv and RvΔB transformants. The transformants were seeded at A₆₀₀ of  ~0.05 in the presence of pristinamycin or IVN as indicated for 5 days and WCLs were resolved on 8% SDS-PAGE and probed with α-PknB and α-GroEL1 antibodies. d Cultures of Rv and RvΔB transformants were initiated at A₆₀₀ of ~0.05 and grown in the presence of 1.5 μg/ml pristinamycin or 0.2 μM IVN as indicated for 6 days. CFUs were evaluated by plating appropriate dilution on 7H11 agar plates containing pristinamycin. Data are representative of the three biologically independent experiments and each experiment was performed in triplicates. e Human monocytic cell line THP-1 was differentiated to macrophages and was infected at 1:10 M.O.I. with Rv and RvΔB transformants grown to A₆₀₀ of ~0.6–1.0 in the presence of pristinamycin. IVN was added in the media to induce the expression of episomal copy and CFUs were enumerated 72 h p.i. Data are representative of one of the two biological replicates and each experiment was performed in triplicates. Data represents mean + SD. Statistical analysis was performed with the unpaired t-test using Graphpad software. ***p < 0.0005. Source data are provided as a Source Data file