Data underlying the figures in the publication “Systematic Engineering of Artificial Metalloenzymes for New-to-Nature Reactions”, published in Sci. Adv., 2021, 7, eabe420RSC. https://advances.sciencemag.org/content/7/4/eabe4208.abstract Table of contents: 1. Dataset 1; Excel file containing the dataset for the Figures in the publication. It contains the activity of 400 artificial metalloenzymes measured for five reactions. In addition, the expression level of all mutants as well as validation experiments in vivo and in vitro are provided. As outlined in the file, some data were processed by subtracting a blank and normalizing to the optical density of the cultures as well as to a wild type control. Mutants are referred to by the amino acids at positions 112 and 121 of the protein.
Pd im aktiven Zentrum: Der Einbau eines biotinylierten Palladiumdiphosphans in Streptavidin ergab ein künstliches Metalloenzym, das asymmetrische allylische Alkylierungen katalysiert (siehe Schema). Chemogenetische Katalysatoroptimierung durch Einführung eines Abstandhalters (roter Stern) zwischen Biotin (grünes Dreieck) und Pd und anschließende Sättigungsmutagenese an der Position S112X ergaben R- und S-selektive asymmetrische allylische Alkylasen.
Abstract In der letzten Dekade fanden künstliche Metalloenzyme einige Aufmerksamkeit als eine mögliche Lösung für ungeklärte Probleme der organischen Synthesechemie. Während gängige Übergangsmetallkatalysatoren meist nur die erste Koordinationssphäre nutzen, um Reaktivität und Selektivität zu kontrollieren, können künstliche Metalloenzyme sowohl die erste als auch die zweite Koordinationssphäre modulieren. Dieser Unterschied manifestiert sich in den teilweise einzigartigen Reaktivitätsprofilen künstlicher Metalloenzyme. Dieser Aufsatz fasst die Versuche zusammen, die zweite Koordinationssphäre künstlicher Metalloenzyme durch genetische Modifizierungen der Proteinsequenz anzupassen. Es werden erfolgreiche Versuche und kreative Wege zur Überwindung aufgetretener Schwierigkeiten hervorgehoben.
Three-legged piano stool complexes are prototypical organometallic complexes relevant to a wide range of chemically relevant questions. Force field parametrization of transition-metal complexes is difficult and underdeveloped, and metal-specific force fields and software are required. Here we report our efforts to derive parameters for the conventional CHARMM and the Valbond-CHARMM force fields for d6-piano stool complexes. In Valbond-CHARMM, the usual angular term is replaced with hybrid orbital strength functions. These functions describe the energy not only of distorted bond angles around the minimum but also at very large distortions. Structure optimizations led to a good agreement between the calculated force field and the X-ray structures. They were comparable to RMSDs obtained between X-ray and DFT structures. In addition, and contrary to treating the systems with DFT, molecular dynamics simulations on the multiple nanosecond time scale are possible and allow to compute meaningful structural and energetic observables. Explicit solvent simulations of the complexes in methanol and water allow to determine the solvent distribution around the complexes. The parametrization presented here will be a useful starting point for dynamics investigations of catalysts in structurally more demanding environments.
Cupredoxins are electron-transfer proteins that have active sites containing a mononuclear Cu center with an unusual trigonal monopyramidal structure (Type 1 Cu). A single Cu-Scys bond is present within the trigonal plane that is responsible for its unique physical properties. We demonstrate that a cysteine-containing variant of streptavidin (Sav) can serve as a protein host to model the structure and properties of Type 1 Cu sites. A series of artificial Cu proteins are described that rely on Sav and a series of biotinylated synthetic Cu complexes. Optical and EPR measurements highlight the presence of a Cu-Scys bond, and XRD analysis provides structural evidence. We further provide evidence that changes in the linker between the biotin and Cu complex within the synthetic constructs allows for small changes in the placement of Cu centers within Sav that have dramatic effects on the structural and physical properties of the resulting artificial metalloproteins. These findings highlight the utility of the biotin-Sav technology as an approach for simulating active sites of metalloproteins.
Abstract Atropisomers – separable conformers that arise from restricted single‐bond rotation – are frequently encountered in medicinal chemistry. However, preparing such compounds with the desired configuration can be challenging. Herein, we present a biocatalytic strategy for achieving atroposelective synthesis relying on artificial metalloenzymes (ArMs). Based on the biotin‐streptavidin technology, we constructed ruthenium‐bearing ArMs capable of producing atropisomeric binaphthalene compounds through ring‐closing metathesis in aqueous media. Further, we show that atroposelectivity can be fine‐tuned by engineering two close‐lying amino acid residues within the streptavidin host protein. The resulting ArMs promote product formation with enantiomeric ratios of up to 81 : 19, while small‐molecule catalysts for atroposelective metathesis under aqueous reaction conditions are yet unknown. This study represents the first demonstration that stereoselective metathesis can be achieved by an artificial metalloenzyme.
