Fig. 3: Crystal structure of MexL15-212 and the key amino acids for DNA-binding. | Nature Communications

Fig. 3: Crystal structure of MexL15-212 and the key amino acids for DNA-binding.

From: Dual-function regulator MexL as a target to control phenazines production and pathogenesis of Pseudomonas aeruginosa

Fig. 3

a Ribbon diagram of the MexL15-212 dimer. Subunit A was shown in green; subunit B was shown in cyans. LBD, ligand-binding domain in C-terminus, consist of seven α helices, α4 to α10. DBD, the N-terminal DNA-binding domain is composed of three helices, α1 to α3. b The alanine (A) of 47, glycine (G) of 48 and tyrosine (Y) of 55 in MexL was highly conserved in TetR family regulator. Shown was the multiple sequence alignment of the HTH DNA-binding domain of MexL and other TetR family regulator, including FadR from Bacillus halodurans, MexZ of P. aeruginosa, MtrR of Neisseria gonorrhoeae, AcrR of Escherichia coli, AmrR of Burkholderia pseudomallei and SmeT of Stenotrophomonas maltophilia. c Structure alignment between MexL and MexL(A47D). MexL and MexL(A47D) are shown in green and magenta, respectively. The hydrogen bond interaction between D47 and R29 is shown as black dashed lines. d Electrophoretic mobility shift assay (EMSA) detection of binding of MexL and MexL(A47D) to the mexJ, phz1, phz2 and phzM promoter, respectively. Representative images from three independent experiments. e PYO production of the PAO1/pUCP20, ΔmexL/pUCP20, ΔmexL/pMexL and ΔmexL/pMexL (A47D) were measured after culture in LB medium for 16 h. (n = 3 independent experiments). f RT-qPCR determination of the expression levels of phzA1, phzA2 and phzM genes in the PAO1/pUCP20, ΔmexL/pUCP20, ΔmexL/pMexL and ΔmexL/pMexL (A47D). (n = 3 independent experiments). P values were determined using two-tailed Student’s t test. Significance was indicated by a P value. ns, non-significant, ***P < 0.001. e P = 0.000945 (PAO1/pUCP20 vs ΔmexL/pUCP20), P = 0.000792 (PAO1/pUCP20 vs ΔmexL/pMexL(A47D)). f P = 0.000161(PAO1/pUCP20 vs ΔmexL/pUCP20 (phzA1)), P = 0.00015 (ΔmexL/pMexL vs ΔmexL/pMexL(A47D) (phzA1)), P = 0.000328 (PAO1/pUCP20 vs ΔmexL/pUCP20 (phzM)), P = 0.00063 (ΔmexL/pMexL vs ΔmexL/pMexL(A47D) (phzM)), P = 0.000349 (PAO1/pUCP20 vs ΔmexL/pUCP20 (phzA2)) and P = 0.000981 (ΔmexL/pMexL vs ΔmexL/pMexL(A47D) (phzA2)).

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