Fig. 4

Molecular mechanisms of MNQ interacting with Tet(X3)/Tet(X4). The catalytic activity of Tet(X3) (a) and Tet(X4) (b) following the diverse concentrations of MNQ incubation. The circular dichroism (CD) spectroscopy analysis of Tet(X3) (c–d) and Tet(X4) (e–f) treated with DMSO or 128 μg/mL MNQ. The binding mode of MNQ/Tet(X3) complex (g) or MNQ/Tet(X4) complex (h) obtained from molecular docking simulations. Decomposition of the binding energy (including E_vdw, E_ele, E_solv, and E_total) on the binding residues between MNQ and Tet(X3) (i) or Tet(X4) (j). The binding constants of MNQ engaging with Tet(X3)-WT and the mutants (k), Tet(X4)-WT and the mutants (l). m–n The synergistic activity of MNQ against Tet(X3)/Tet(X4)-mutated stains at the indicated binding sites was determined by broth micro-dilution methods. o Tet(X3)/Tet(X4) levels in the precipitates of bacterial cultures treated with or without MNQ were assessed by western blotting. p The mRNA levels of tet(X3)/tet(X4) in the E. coli DH5α + pAM401-tet(X3)/tet(X4) were quantified by real-time PCR and normalized to 16S rRNA. The data are representative of three biological replicates and expressed as ± SEM. **P < 0.01