Development of a bacterial biosensor for nitrotoluenes: the crystal structure of the transcriptional regulator DntR.
The transcriptional regulator DntR, a member of the LysR family, is a central element in a prototype bacterial cell-based biosensor for the detection of hazardous contamination of soil and groundwater by dinitrotoluenes. To optimise the sensitivity of the biosensor for such compounds we have chosen a rational design of the inducer-binding cavity based on knowledge of the three-dimensional structure of DntR. We report two crystal structures of DntR with acetate (resolution 2.6 angstroms) and thiocyanate (resolution 2.3 angstroms), respectively, occupying the inducer-binding cavity. These structures allow for the construction of models of DntR in complex with salicylate (Kd approximately or = 4 microM) and 2,4-dinitrotoluene that provide a basis for the design of mutant DntR with enhanced specificity for dinitrotoluenes. In both crystal structures DntR crystallises as a homodimer with a "head-to-tail" arrangement of monomers in the asymmetric unit. Analysis of the crystal structure has allowed the building of a full-length model of DntR in its biologically active homotetrameric form consisting of two "head-to-head" dimers. The implications of this model for the mechanism of transcription regulation by LysR proteins are discussed.
Smirnova
Irina A
IA
Department of Biochemistry and Biophysics, Arrhenius Laboratories for Natural Sciences, Stockholm University, SE-106 91 Stockholm, Sweden.
Dian
Cyril
C
Leonard
Gordon A
GA
McSweeney
Seán
S
Birse
Darcy
D
Brzezinski
Peter
P
D001483:Base Sequence; D015374:Biosensing Techniques; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D017931:DNA Primers; D004136:Dinitrobenzenes / Q000032:analysis*; D024202:Electrophoretic Mobility Shift Assay; D008958:Models, Molecular; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics
D017931:DNA Primers; D004136:Dinitrobenzenes; D014157:Transcription Factors; C016403:2,4-dinitrotoluene