Thaxtomins, a family of phytotoxins produced by Streptomyces spp., can cause dramatic plant cell hypertrophy and seedling stunting. Thaxtomin A is the dominant form from Streptomyces scabies and has demonstrated herbicidal action. TxtE, a cytochrome P450 enzyme from Streptomyces scabies 87.22, catalyzes direct nitration of the indolyl moiety of L-tryptophan to L-4-nitrotryptophan using nitric oxide, dioxygen and NADPH. The crystal structure of TxtE was determined at 2.1 Å resolution and described in this work. A clearly defined substrate access channel is observed and can be classified as channel 2a, which is common in bacteria cytochrome P450 enzymes. A continuous hydrogen bond chain from the active site to the external solvent is observed. Compared with other cytochrome P450 enzymes, TxtE shows a unique proton transfer pathway which crosses the helix I distortion. Polar contacts of Arg59, Tyr89, Asn293, Thr296, and Glu394 with L-tryptophan are seen using molecular docking analysis, which are potentially important for substrate recognition and binding. After mutating Arg59, Asn293, Thr296 or Glu394 to leucine, the substrate binding ability of TxtE was lost or decreased significantly. Based on the docking and mutation results, a possible mechanism for substrate recognition and binding is proposed.

VqmA is a highly conserved transcriptional regulator of the quorum sensing system of Vibrio cholerae, a major human pathogen that continues to imperil human health. VqmA represses biofilm formation and plays an important role in V. cholerae pathogenicity in the human host. Despite the good understanding of VqmA’s biological function, the molecular mechanisms by which its specific ligand (and effector), 3,5-dimethylpyrazine-2-ol (DPO), controls transcription of the target gene, vqmR, remain obscure. To elucidate the molecular mechanism of DPO binding, we used structural analyses and biochemical assays to study the V. cholerae VqmA–DPO–DNA complex. These analyses revealed that VqmA contains an N-terminal homodimer domain (PAS) and a C-terminal DNA binding domain (DBD). We observed that VqmA directly binds to a DPO molecule via a compact hydrophobic pocket, consisting of a six-stranded antiparallel β-sheet and several α-helices. We also found that the VqmA dimer interacts with the quasi-palindromic sequence of the vqmR promoter through its DBD. The results of the biochemical studies indicated that a water atom and the VqmA residues Phe67 and Lys101 play a key role in effector recognition, which is also assisted by Tyr36 and Phe99. This is the first molecular-level view of the VqmA dimer bound to DPO and DNA. The structure–function analyses presented here improve our understanding of the complex mechanisms in the transcriptional regulation of VqmA in Vibrio spp. and may inform the design of drugs to manage V. cholerae infections.