2XDN

Transcription factor TtgR H67A mutant

2XDN の概要

• エントリーDOI: 10.2210/pdb2xdn/pdb
• 関連するPDBエントリー: 2UXH 2UXI 2UXO 2UXP 2UXU
• 分子名称: HTH-TYPE TRANSCRIPTIONAL REGULATOR TTGR (2 entities in total)
• 機能のキーワード: transcription, transcription regulation, tetr family
• 由来する生物種: PSEUDOMONAS PUTIDA
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 95293.34

構造登録者

Daniels, C.,Lu, D.,Zhang, X.,Ramos, J.L. (登録日: 2010-05-04, 公開日: 2010-05-12, 最終更新日: 2024-05-08)

主引用文献

Daniels, C.,Daddaoua, A.,Lu, D.,Zhang, X.,Ramos, J.L.
Domain Cross-Talk During Effector Binding to the Multidrug Binding Ttgr Regulator.
J.Biol.Chem., 285:21372-, 2010

PubMed Abstract: A major mechanism of antibiotic resistance in bacteria is the active extrusion of toxic compounds through membrane-bound efflux pumps. The TtgR protein represses transcription of ttgABC, a key efflux pump in Pseudomonas putida DOT-T1E capable of extruding antibiotics, solvents, and flavonoids. TtgR contains two distinct and overlapping ligand binding sites, one is broad and contains mainly hydrophobic residues, whereas the second is deep and contains polar residues. Mutants in the ligand binding pockets were generated and characterized using electrophoretic mobility shift assays, isothermal titration calorimetry, and promoter expression. Several mutants were affected in their response to effectors in vitro: mutants H70A, H72A, and R75A did not dissociate from promoter DNA in the presence of chloramphenicol. Other mutants exhibited altered binding to the operator: L66A and L66AV96A mutants bound 3- and 15-fold better than the native protein, whereas the H67A mutant bound with 3-fold lower affinity. In vivo expression assays using a fusion of the promoter of ttgA to lacZ and antibiotic tolerance correlated with the in vitro observations, namely that mutant H67A leads to increased basal expression levels and enhances antibiotic tolerance, whereas mutants L66A and L66AV96A exhibit lower basal expression levels and decreased resistance to antibiotics. The crystal structure of TtgR H67A was resolved. The data provide evidence for the inter-domain communication that is predicted to be required for the transmission of the effector binding signal to the DNA binding domain and provide important information to understand TtgR/DNA/effector interactions.

PubMed: 20435893
DOI: 10.1074/JBC.M110.113282

実験手法

X-RAY DIFFRACTION (2.2 Å)