title: How does Mg(2+) affect the binding of anhydrotetracycline in the TetR protein?
abstract: The binding of anhydrotetracycline (atc) in wild-type TetR(D), TetR(B), and four single tryptophan mutants of TetR(B) was investigated by UV/vis absorption, steady state, and time-resolved fluorescence spectroscopy. From absorption titration experiments with Mg2+, we conclude that binding of one [atc-Mg]+ complex in the homodimer causes changes in the protein conformation around the second binding pocket. In the presence of absence of Mg2+, several different groups of atc-protein arrangements must exist, each with a characteristic atc fluorescence decay time. Taking into account the results of molecular dynamics (MD) simulations, we propose as one possible origin for such a differentiation teh extent of hydrogen bonding between atc and the surrounding amino acids. Binding of Mg2+ should change the arrangement of the surrounding amino acids such that some of the excited atc molecules do not undergo the relaxation process typical for free atc. The MD simulations also show that the pattern of intra- and intermolecular hydrogen bonding in the two monomeric units is no correlated, thereby leading to different fluorescence kinetics for atc in the two monomeric units. Furthermore, it is suggested that hydrogen bonding between Arg104 and O10 of anhydrotetracycline could regulate the relaxation processes of excited anhydrotetracycline.
authors.affiliation: Institut für Physikalische und Theiretische Chemie, Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Clemens F
authors.initials: CF
authors.lastname: Leypold
authors.forename: Dan-Thomas
authors.initials: DT
authors.lastname: Marian
authors.forename: Ciprian
authors.initials: C
authors.lastname: Roman
authors.forename: Siegfried
authors.initials: S
authors.lastname: Schneider
authors.forename: Peter
authors.initials: P
authors.lastname: Schubert
authors.forename: Oliver
authors.initials: O
authors.lastname: Scholz
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Hillen
authors.forename: Timothy
authors.initials: T
authors.lastname: Clark
authors.forename: Harald
authors.initials: H
authors.lastname: Lanig
mesh_terms: D001426:Bacterial Proteins / Q000187:drug effects / Q000378:metabolism*; D004492:Edetic Acid; D008274:Magnesium / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry; D013754:Tetracyclines / Q000737:chemistry*; D015534:Trans-Activators / Q000187:drug effects / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; C116102:TetR protein, Clostridium tetani; D013754:Tetracyclines; D015534:Trans-Activators; C016229:4-epianhydrotetracycline; D004492:Edetic Acid; D008274:Magnesium