Structural, kinetic, and docking studies of artificial imine reductases based on biotin-streptavidin technology: An induced lock-and-key hypothesis

Victor Muñoz Robles, Marc Dürrenberger, Tillmann Heinisch, Agustí Lledós, Tilman Schirmer, Thomas R. Ward, Jean Didier Maréchal

Research output: Contribution to journalArticleResearchpeer-review

47 Citations (Scopus)

Abstract

© 2014 American Chemical Society. An artificial imine reductase results upon incorporation of a biotinylated Cp Ir moiety (Cp∗ = C<inf>5</inf>Me<inf>5</inf><sup>-</sup>) within homotetrameric streptavidin (Sav) (referred to as CpIr(Biot-p-L)Cl] ⊂ Sav). Mutation of S112 reveals a marked effect of the Ir/streptavidin ratio on both the saturation kinetics as well as the enantioselectivity for the production of salsolidine. For [CpIr(Biot-p-L)Cl] ⊂ S112A Sav, both the reaction rate and the selectivity (up to 96% ee (R)-salsolidine, k<inf>cat</inf> 14-4 min<sup>-1</sup> vs [Ir], K<inf>M</inf> 65-370 mM) decrease upon fully saturating all biotin binding sites (the ee varying between 96% ee and 45% ee R). In contrast, for [CpIr(Biot-p-L)Cl] ⊂ S112K Sav, both the rate and the selectivity remain nearly constant upon varying the Ir/streptavidin ratio [up to 78% ee (S)-salsolidine, k<inf>cat</inf> 2.6 min<sup>-1</sup>, K<inf>M</inf> 95 mM]. X-ray analysis complemented with docking studies highlight a marked preference of the S112A and S112K Sav mutants for the S<inf>Ir</inf> and R<inf>Ir</inf> enantiomeric forms of the cofactor, respectively. Combining both docking and saturation kinetic studies led to the formulation of an enantioselection mechanism relying on an "induced lock-and-key" hypothesis: the host protein dictates the configuration of the biotinylated Ir-cofactor which, in turn, by and large determines the enantioselectivity of the imine reductase.
Original languageEnglish
Pages (from-to)15676-15683
JournalJournal of the American Chemical Society
Volume136
Issue number44
DOIs
Publication statusPublished - 5 Nov 2014

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