Tetracycline Analogs Affecting Binding to Tn10-Encoded Tet Repressor Trigger the Same Mechanism of Induction

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• 作者：Thomas Lederer ; Martin Kintrup ; Masayuki Takahashi ; Phaik-Eng Sum ; George A. Ellestad ; and Wolfgang Hillen
• 刊名：Biochemistry
• 出版年：1996
• 出版时间：June 11, 1996
• 年：1996
• 卷：35
• 期：23
• 页码：7439 - 7446
• 全文大小：349K
• 年卷期：v.35,no.23(June 11, 1996)
• ISSN：1520-4995

文摘

We examined the influence of substituents in tetracycline (tc)analogs modified at positions 2and 4-9 and anhydrotetracycline (atc) on induction of theTn10-encoded Tet repressor (TetR) by aquantitative in vitro induction assay. The equilibriumassociation constants of the modified tc to TetRwere independently determined to distinguish effects on binding fromthose on induction. We found acorrelation between the binding affinity and induction of TetR for mosttc analogs. While a substitutionat position 5 revealed only minor effects, changes at position 6increased binding and induction efficienciesup to 20-fold. A chlorine at position 7 or 8 enhanced binding andinduction about 4- and 9-fold,respectively. Substituents at position 9 decreased binding up to5-fold. Epimerization of the dimethylaminofunction at position 4 in 4-epi-tc resulted in about 300-fold-reducedbinding and 80-fold-reduced induction.Substitution of this grouping by hydrogen in4-de(dimethylamino)-tc resulted in no binding and noinduction.The respective atc analog failed to induce as well, althoughbinding was still observed. The dimethylaminofunction may, thus, play a role in triggering the conformational changeof TetR necessary for induction.Substitution of the 2-carboxamido by a nitrilo function did notinfluence binding and induction efficiencies.Atc showed about 30-fold increased binding and induction, beingthe most effective inducer tested in thisstudy. The equilibrium association constants of mostTetR-[Mg-tc]⁺ andTetR-([Mg-tc]⁺)₂ analogcomplexes with tet operator are decreased about10²- and 10⁸-fold, respectively, as compared tothose offree TetR. This suggests that these tc analogs share the samemolecular mechanism of TetR induction.