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Protonation Patterns in Tetracycline:Tet Repressor Recognition: Simulations and Experiments
Author(s) -
Aleksandrov Alexey,
Proft Juliane,
Hinrichs Winfried,
Simonson Thomas
Publication year - 2007
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.200600535
Subject(s) - tetr , protonation , chemistry , ligand (biochemistry) , repressor , amide , molecular dynamics , crystallography , tetracycline , stereochemistry , computational chemistry , ion , biochemistry , organic chemistry , gene , gene expression , receptor , antibiotics
Resistance to the antibiotic tetracycline (Tc) is regulated by its binding as a Tc:Mg 2+ complex to the Tet Repressor protein (TetR). Tc:TetR recognition is a complex problem, with the protein and ligand each having several possible conformations and protonation states, which are difficult to elucidate by experiment alone. We used a combination of free‐energy simulations and crystallographic analysis to investigate the electrostatic interactions between protein and ligand and the possible role of induced fit in Tc binding. Tc in solution was described quantum mechanically, while Tc:TetR interactions were described by a recent, high‐quality molecular‐mechanics model. The orientations of the amide and imidazole groups were determined experimentally by a careful analysis of Debye–Waller factors in alternate crystallographic models. The agreement with experiment for these orientations suggested that the simulations and their more detailed, thermodynamic predictions were reliable. We found that the ligand prefers an extended, zwitterionic state both in solution and in complexation with the protein. Tc is thus preorganized for binding, while the protein combines lock‐and‐key behavior for regions close to the ligand's amide, enolate, and ammonium groups, with an induced fit for regions close to the Mg 2+ ion. These insights and the modeling techniques employed should be of interest for engineering improved TetR ligands and improved TetR proteins for gene regulation, as well as for drug design.