Robles, Victor Muñoz Dürrenberger, Marc Heinisch, Tillmann Lledós, Agustí Schirmer, Tilman Ward, Thomas R. Maréchal, Jean-Didier Structural, Kinetic, and Docking Studies of Artificial Imine Reductases Based on Biotin–Streptavidin Technology: An Induced Lock-and-Key Hypothesis An artificial imine reductase results upon incorporation of a biotinylated Cp*Ir moiety (Cp* = C<sub>5</sub>Me<sub>5</sub><sup>–</sup>) within homotetrameric streptavidin (Sav) (referred to as Cp*Ir­(Biot-<i>p</i>-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 [Cp*Ir­(Biot-<i>p</i>-L)­Cl] ⊂ S112A Sav, both the reaction rate and the selectivity (up to 96% ee (<i>R</i>)-salsolidine, <i>k</i><sub>cat</sub> 14–4 min<sup>–1</sup> vs [Ir], <i>K</i><sub>M</sub> 65–370 mM) decrease upon fully saturating all biotin binding sites (the ee varying between 96% ee and 45% ee <i>R</i>). In contrast, for [Cp*Ir­(Biot-<i>p</i>-L)­Cl] ⊂ S112K Sav, both the rate and the selectivity remain nearly constant upon varying the Ir/streptavidin ratio [up to 78% ee (<i>S</i>)-salsolidine, <i>k</i><sub>cat</sub> 2.6 min<sup>–1</sup>, <i>K</i><sub>M</sub> 95 mM]. X-ray analysis complemented with docking studies highlight a marked preference of the S112A and S112K Sav mutants for the <i>S</i><sub>Ir</sub> and <i>R</i><sub>Ir</sub> 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. enantioselectivity;RIr enantiomeric forms;Docking Studies;enantioselection mechanism;ee;selectivity;docking studies;S 112;saturation kinetics;imine reductase results;kcat;S 112K Sav mutants;Artificial Imine Reductases;mM;reaction rate;KM;Cp;C 5Me;S 112A;biotinylated;homotetrameric streptavidin;imine reductase;biotin binding sites;host protein dictates 2014-11-05
    https://acs.figshare.com/articles/journal_contribution/Structural_Kinetic_and_Docking_Studies_of_Artificial_Imine_Reductases_Based_on_Biotin_Streptavidin_Technology_An_Induced_Lock_and_Key_Hypothesis/2239477
10.1021/ja508258t.s001