title: Tetracycline analogs affecting binding to Tn10-Encoded Tet repressor trigger the same mechanism of induction.
abstract: We examined the influence of substituents in tetracycline (tc) analogs modified at positions 2 and 4-9 and anhydrotetracycline (atc) on induction of the Tn10-encoded Tet repressor (TetR) by a quantitative in vitro induction assay. The equilibrium association constants of the modified tc to TetR were independently determined to distinguish effects on binding from those on induction. We found a correlation between the binding affinity and induction of TetR for most tc analogs. While a substitution at position 5 revealed only minor effects, changes at position 6 increased binding and induction efficiencies up to 20-fold. A chlorine at position 7 or 8 enhanced binding and induction about 4- and 9-fold, respectively. Substituents at position 9 decreased binding up to 5-fold. Epimerization of the dimethylamino function at position 4 in 4-epi-tc resulted in about 300-fold-reduced binding and 80-fold-reduced induction. Substitution of this grouping by hydrogen in 4-de(dimethylamino)-tc resulted in no binding and no induction. The respective atc analog failed to induce as well, although binding was still observed. The dimethylamino function may, thus, play a role in triggering the conformational change of TetR necessary for induction. Substitution of the 2-carboxamido by a nitrilo function did not influence binding and induction efficiencies. Atc showed about 30-fold increased binding and induction, being the most effective inducer tested in this study. The equilibrium association constants of most TetR-[Mg-tc]+ and TetR-([Mg-tc]+)2 analog complexes with tet operator are decreased about 10(2)- and 10(8)-fold, respectively, as compared to those of free TetR. This suggests that these tc analogs share the same molecular mechanism of TetR induction.
authors.affiliation: Lehrstuhl für Mikrobiologie, Institut für Mikrobiologie, Bioachemie und Genetik der Friedrich-Alexander-universität Erlangen-Nürnberg, FRG.
authors.forename: T
authors.initials: T
authors.lastname: Lederer
authors.forename: M
authors.initials: M
authors.lastname: Kintrup
authors.forename: M
authors.initials: M
authors.lastname: Takahashi
authors.forename: P E
authors.initials: PE
authors.lastname: Sum
authors.forename: G A
authors.initials: GA
authors.lastname: Ellestad
authors.forename: W
authors.initials: W
authors.lastname: Hillen
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism; D001665:Binding Sites; D002352:Carrier Proteins; D004251:DNA Transposable Elements / Q000187:drug effects*; D006860:Hydrogen Bonding; D007700:Kinetics; D008274:Magnesium; D015394:Molecular Structure; D010957:Plasmids; D012097:Repressor Proteins / Q000737:chemistry / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013754:Tetracyclines / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D004251:DNA Transposable Elements; D012097:Repressor Proteins; C083139:Tet O resistance protein, Bacteria; D013754:Tetracyclines; C034811:tetracycline resistance-encoding transposon repressor protein; D008274:Magnesium