Supplementary MaterialsTable_1. The results of molecular mechanics generalized-Born surface area calculations indicate that the binding free energy of rifampicin with three mutants decreases. In addition, the dynamic network analysis and residue interaction network analysis show that when H451 was K-Ras G12C-IN-2 mutated, the interactions of residue 451 with its adjacent residues such as Q438, F439, M440, D441, and S447 disappeared or weakened, increasing the flexibility of binding pocket. At the same time, the disappearance of hydrogen bonds between R613 and rifampicin caused by the flipping of R613 is another important reason for the reduction of binding ability CSP-B of rifampicin in H451D/Y mutants. In H451R mutant, the mutation causes the binding pocket change too much so that the position of rifampicin has K-Ras G12C-IN-2 a large movement in the binding pocket. In this study, the resistance mechanism of rifampicin at the atomic level is proposed. The proposed drug-resistance mechanism will provide the valuable guidance for the design of antituberculosis drugs. (Mtb), is the leading cause from a single infectious agent world-wide. Mtb, a pathogenic bacterium varieties of the family members RNA polymerase (Mtb-RNAP). A lot K-Ras G12C-IN-2 more than 95% from the rifampin-resistant strains possess mutations in a little region described rifampicin resistance-determining area in Mtb-RNAP (Morlock et al., 2000; Zaw et al., 2018). The most frequent mutation in rifampicin resistance-determining area are S456, H451, and D441, related to S531, H526, and D516 in activity test of rifampicin by Bodmer et al. (1995) have been proven that H451D/Y/R mutations might lead to high-level level of resistance to rifampicin. After a lot more than two K-Ras G12C-IN-2 decades, the particular level and frequency of resistance to rifampicin are increasing also. In 2017, there is about 558,000 new cases of rifampicin-resistant tuberculosis (RR-TB), of which 82% are MDR-TB and about 230,000 deaths from MDR/RR-TB (Organization, 2018). Currently, although MDR/RR-TB can be cured with the second-line drugs (e.g., fluoroquinolone and an injectable aminoglycoside), poor efficiency, high toxicity, and expensive price of these drugs make it still difficult for many MDR-TB patients. In some cases, more severe extensively drug-resistant TB may occur, and it will not respond to the most effective second-line anti-TB drugs (Sotgiu et al., 2015; Jeon, 2017; Tiberi et al., 2018). Obviously, the development of new anti-TB drugs is urgent, and exploring the resistance mechanism of rifampicin is of great significance for the discovery of effective drugs. In this work, in order to uncover the resistance mechanism of Mtb to rifampicin due to the mutation of Mtb-RNAP at position 451, three independent molecular dynamics (MD) simulations for the wild-type Mtb-RNAP and H451D/Y/R mutants were carried out. Based on the obtained trajectories, the molecular mechanics generalized-Born surface area (MM-GBSA) method was applied to calculate the binding free energy of rifampicin with Mtb-RNAP. Furthermore, dynamic network analysis combined with residue interaction network (RIN) analysis was used to show the detailed changes of interactions among the residues surrounding the binding pocket. With the structural and energy analysis, a possible rifampicin-resistant mechanism was also proposed. Compared with the traditional experimental method, MD simulations can show the intuitive and dynamics interaction change process between rifampicin and Mtb-RNAP due to the point mutation. Together with the energy analysis and the dynamics network analysis, the present study show the essential reason of Mtb-RNAP resistant to rifampicin, which can provide the useful guidance for the further drug design against drug resistance. Materials and Methods Systems Preparation The initial atomic coordinate of the wild-type Mtb-RNAP with rifampicin was obtained from Protein Data Bank (Protein Data Bank ID: 5UHB). The crystal structure of Mtb-RNAP reported by Lin et al. (2017) reveals that Mtb-RNAP is composed of six chains, for the A, B chains encoded by the rpoA gene, the C chain encoded from the rpoB gene (Miller et al., K-Ras G12C-IN-2 1994), as well as the D, E, F stores encoded by rpoC, rpoZ, and rpoD, respectively. Rifampicin binds in the energetic site from the C string (demonstrated in Shape 1) and inhibits the DNA-directed RNA synthesis of Mtb (McClure and Cech, 1978; Campbell et al., 2001; Somoskovi et al., 2001). Due to the fact the acceleration to simulate the complete Mtb-RNAP (~3,826 residues) can be too slow, just the C string complexed with rifampicin was used and extracted mainly because the original structure of simulations. Furthermore, the deletion of additional stores shall make the residues from the user interface between your two stores unpredictable, which can be inconsistent with this in the multimer. Therefore, to simulate the constant state of user interface in the multimer, some relatively versatile and definately not the energetic site amino acidity residues were erased. The three-dimensional constructions of three mutants (H451D/Y/R) had been acquired by mutating H451 residue in crazy type. Open up in.