Molecular modelling and competitive inhibition of a Mycobacterium tuberculosis multidrug-resistance efflux pump.

Tuberculosis is a major cause of mortality and morbidity in developing countries, and the emergency of multidrug and extensive drug resistance cases is an utmost issue on the control of the disease. Despite the efforts on the development of new antibiotics, eventually there will be strains resistant to them as well. Efflux plays an important role in the evolution of resistance in Mycobacterium tuberculosis. Tap is an important efflux pump associated with tuberculosis resistant to isoniazid, rifampicine and ofloxacin and with multidrug resistance. The development of efflux inhibitors for Tap could raise the effectiveness of second line drugs and reduce the duration of the current treatment. Therefore the objective of this study is to build a reliable molecular model of Tap efflux pump and test the possible competitive inhibition between efflux inhibitors and antibiotics in the optimized structure. We built twenty five Tap models with molecular modelling to elect the best according to the results of the validation analysis. The elected model went through to a 100 ns molecular dynamics simulation in a lipid bilayer, and the resulting optimized structure was used in docking studies to test if the used efflux inhibitors may act via competitive inhibition on antibiotics. The validation results pointed the model built by Phyre2 as the closest to a possible native Tap structure, and therefore it was the elected model. RSMD analysis revealed the model is stable, where the predicted binding site stabilized between 15 and 20 ns, maintaining the RMSD at around 0.35 Å throughout the molecular dynamics simulation in a lipid bilayer. Therefore this model is reliable and can also be used for further studies. The docking studies showed a possibility of competitive inhibition by NUNL02 on ofloxacin and bedaquiline, and by verapamil on ofloxacin and rifampicin. This presents the possibility that NUNL02 and verapamil are possible inhibitors of Tap efflux and highlights the importance of including efflux inhibitors as adjuvants to the tuberculosis therapy, as it indicates a possible extrusion of ofloxacin, rifampicin and bedaquilin by Tap.