DNA binding exerted by a bacterial gene regulator with an extensive coiled-coil domain.
Although quite common in the eukaryotic cell, bacterial proteins with an extensive coiled-coil domain are still relatively rare. One of the few thus far documented examples, TlpA from Salmonella typhimurium, is characterized by a remarkably long (250 amino acids) alpha-helical coiled-coil domain. Herein, we demonstrate that TlpA is a novel sequence-specific DNA-binding protein. Several tlpA deletion mutants have been constructed, and their corresponding protein products were purified and tested for DNA binding. Two of the mutant proteins were shown to be deficient in DNA binding. Both mutants were analyzed by circular dichroism and electron microscopy, supporting the notion that mutant proteins wre shown to be deficient in DNA binding. Both mutants were analyzed by circular dichroism and electron microscopy, supporting the notion that mutant proteins were largely intact despite lacking the amino acid residues necessary for DNA binding. In vivo studies with transcriptional tlpA-lacZ fusions demonstrated that TlpA acts as a repressor. Using the repressor phenotype as a readout, the chain exchange previously described in vitro could also be confirmed in vivo. We believe the coiled-coil domain acts not only as a dimerization interface but could also serve a role as a flexible modulator of the protein-DNA interaction.
Hurme
R
R
Microbiology and Tumor Biology Center, Karolinska Institute, Stockholm, Sweden. reini.hurme@mtc.ki.se
Berndt
K D
KD
Namork
E
E
Rhen
M
M
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011487:Protein Conformation; D012486:Salmonella typhimurium / Q000378:metabolism*; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C075138:TlpA protein, Salmonella; D004247:DNA