W gradient in the N terminus (blue) to the C terminus (red). b, ribbon diagram of the Rv0678 dimer. Each subunit of Rv0678 is labeled using a diverse color (yellow and orange). The bound 2-stearoylglycerol within the dimer is shown in sphere kind (gray, carbon; red, oxygen). The figure was prepared utilizing PyMOL.FIGURE 4. Rigid body rotation of your DNA-binding domain of Rv0678. This is a schematic representation illustrating the conformational change of Rv0678 between the ligand-bound and -unbound structures. Helices 4 and four with the DNA-binding domain are indicated. The ligand is colored blue.As a member with the MarR family members of regulators, the DNAbinding domain of Rv0678 characteristics a Nav1.3 Inhibitor Compound common winged helix-turnhelix binding motif. The two anti-parallel 1 and two strands are found to generate a -hairpin structure, which also types the wing in the DNA-binding domain. The crystal structure with the OhrR-DNA complicated (36) showed that this -hairpin directly participates to speak to the double-stranded DNA and is criticalJUNE six, 2014 ?VOLUME 289 ?NUMBERfor repressor-operator interactions. Yet another vital element from the winged helix-turn-helix motif for DNA recognition is helix 4. Inside the OhrR-DNA complex (36), the Met Inhibitor Molecular Weight corresponding -helix is identified to bind inside the deep major groove from the B-DNA. Protein sequence alignment suggests that Rv0678 consists of three conserved amino acids frequent among members on the MarR household. These three residues, Arg-84,JOURNAL OF BIOLOGICAL CHEMISTRYStructure of your Transcriptional Regulator RvFIGURE 5. Simulated annealing electron density maps plus the 2-stearoylglycerol binding website. a, stereo view of your simulated annealing electron density map in the bound 2-stearoylglycerol within the Rv0678 dimer (the orientation corresponds to the side view of Fig. 1b). The bound 2-stearoylglycerol is shown as a stick model (green, carbon; red, oxygen). The simulated annealing 2Fo Fc electron density map is contoured at 1.2 (blue mesh). The left and proper subunits of Rv0678 are shown as orange and yellow ribbons. b, the 2-stearoylglycerol binding internet site. Amino acid residues inside 3.9 ?in the bound 2-stearoylglycerol (green, carbon; red, oxygen) are shown with one-letter codes. The side chains of selected residues in the right subunit of Rv0678 in Fig. 1b are shown as yellow sticks (yellow, carbon; blue, nitrogen; red, oxygen). Residues in the next subunit of Rv0678 are shown as orange sticks (orange, carbon; blue, nitrogen; red, oxygen). c, schematic representation on the Rv0678 and 2-stearoylglycerol interactions. Amino acid residues within 4.five ?in the bound 2-stearoylglycerol are shown with one-letter codes. Dotted lines, hydrogen bonds. The hydrogen-bonded distances are also indicated.16532 JOURNAL OF BIOLOGICAL CHEMISTRYVOLUME 289 ?Quantity 23 ?JUNE 6,Structure of the Transcriptional Regulator RvFIGURE 6. Identification of the fortuitous ligand by GC-MS. a, electron ionization spectrum from the strongest GC peak at 14.45 min. b, GC-MS spectrum of octadecanoic acid, 2-hydroxyl-1-(hydroxymethyl)ethyl ester in the internal GC-MS library. The ligand was identified as 2-stearoylglycerol.JUNE 6, 2014 ?VOLUME 289 ?NUMBERJOURNAL OF BIOLOGICAL CHEMISTRYStructure of the Transcriptional Regulator RvTABLE 4 Rv0678-ligand contactsContacts inside four.5 ?are listed.Residue-ligand contacts Arg-32 Gln-78 Phe-79 Glu-108 Arg-109 Arg-111 Ala-112 Met-113 Glu-115 Leu-116 Leu-144 Leu-145 Tyr-28 Phe-29 Arg-32 Leu-34 Phe-79 Phe-81 Phe-102 Ala-103 Gly-105.