Detailed studies of the binding mechanism of the Sinorhizobium meliloti transcriptional activator ExpG to DNA.
The exopolysaccharide galactoglucan promotes the establishment of symbiosis between the nitrogen-fixing Gram-negative soil bacterium Sinorhizobium meliloti 2011 and its host plant alfalfa. The transcriptional regulator ExpG activates expression of galactoglucan biosynthesis genes by direct binding to the expA1, expG/expD1 and expE1 promoter regions. ExpG is a member of the MarR family of regulatory proteins. Analysis of target sequences of an ExpG(His)(6) fusion protein in the exp promoter regions resulted in the identification of a binding site composed of a conserved palindromic region and two associated sequence motifs. Association and dissociation kinetics of the specific binding of ExpG(His)(6) to this binding site were characterized by standard biochemical methods and by single-molecule spectroscopy based on the atomic force microscope (AFM). Dynamic force spectroscopy indicated a distinct difference in the kinetics between the wild-type binding sequence and two mutated binding sites, leading to a closer understanding of the ExpG-DNA interaction.
Baumgarth
Birgit
B
Lehrstuhl für Genetik, Fakultät für Biologie, Universität Bielefeld, 33615 Bielefeld, Germany.
Bartels
Frank Wilco
FW
Experimentelle Biophysik, Fakultät für Physik, Universität Bielefeld, 33615 Bielefeld, Germany.
Anselmetti
Dario
D
Experimentelle Biophysik, Fakultät für Physik, Universität Bielefeld, 33615 Bielefeld, Germany.
Becker
Anke
A
Lehrstuhl für Genetik, Fakultät für Biologie, Universität Bielefeld, 33615 Bielefeld, Germany.
Ros
Robert
R
Experimentelle Biophysik, Fakultät für Physik, Universität Bielefeld, 33615 Bielefeld, Germany.
D001483:Base Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D004269:DNA, Bacterial / Q000378:metabolism*; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005685:Galactans / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D005936:Glucans / Q000378:metabolism; D007700:Kinetics; D000455:Medicago sativa; D018625:Microscopy, Atomic Force; D008969:Molecular Sequence Data; D011135:Polysaccharides, Bacterial / Q000378:metabolism; D011401:Promoter Regions, Genetic; D016962:Sinorhizobium meliloti / Q000235:genetics / Q000378:metabolism*; D013559:Symbiosis; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic
D004269:DNA, Bacterial; C100020:EXPG protein, Rhizobium meliloti; D005656:Fungal Proteins; D005685:Galactans; D005936:Glucans; D011135:Polysaccharides, Bacterial; D015534:Trans-Activators; C093768:galactoglucan