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Artificial oxidases and oxygenases for commodity chemistry

Published on 6 July 2020
Dr. Caroline Marchi Delapierre
Associate Professor (HC – HDR) Université Grenoble-Alpes
Laboratoire Chimie et Biologie des Métaux
17 avenue des Martyrs
38 054 Grenoble Cedex 09
Tel : 33 (0)4 38 78 91 04
Fax : 33 (0)4 38 78 91 24

Artificial oxidases and oxygenases for commodity chemistry
S. Ménage and C. Marchi-Delapierre

The field of catalysis needs to combine biocatalysis and inorganic catalysis in order to extend the repertoire of reactions to unnatural ones. This strategy relies into the modification of proteins (or enzymes) to gain unnatural abiotic activities. Among the different approaches, the insertion of an inorganic complex playing the role of the active site into the protein controlling the selectivity of the reaction has been developed in the BioCE team, providing a new functionalization via the protein scaffold.

Several achievements have led to the rationale design of such biohybrids the so called artificial metalloenzymes (ArMs). Using NikA, an E. coli nickel transport protein, efficient catalysis for sulfoxidation (TON up to 600), hydroxychlorination and aromatic hydroxylation have been performed in solution. For example, supramolecular interactions of the protein scaffold with a ruthenium complex led to its activation via a ligand exchange and a flip of the metal spin state, crucial for oxidant activation (collaboration N. Burzlaff) [Lopez et al. Chem. Commun. 2017]. Another strategy of anchoring the complex, called the Trojan horse approach, was used to develop an iron oxidase capable of oxygen transfer thanks to the use of the human serum albumin protein, more efficient than the catalyst alone thanks to a better stabilization of the iron complex toward ligand oxidative degradation [Rondot et al. J. Mol. Catal. A 2016]. Moreover, a rationale of the design of artificial enzymes has been validated with the use of docking calculations in order to select the appropriate substrate [Esmieu et al. Ang. Chem. Int. Ed., 2013].
Despite our effort, the stability of the biohybrids needs to be improved. To ensure their relevance for industry, enzyme immobilization is often a pre-requisite to prevent catalyst degradation and maximizing reusability. While in Nature, intracellular processes spatially and temporally control enzyme availability, chemists have developed catalyst heterogeneization strategies by surface grafting, embedment in gels and crystalline solids using cross-linked enzyme crystals (CLEC). Thanks to the high solvent content present in protein crystals (about 50 %), the CLEC technology may afford the advantage of microporous materials such as zeolites or MCM silica. Our capacity, in collaboration with the team BioCat for the mastering of protein crystallization, urged us to design solid crystals containing our inorganic catalysts. Their catalytic properties were outstanding by the number of turnover measured in the solid (TON above 30 000) while only a TON of 400 was reached in solution. These properties were also observed to a lesser extent during the oxidation of thioethers into sulfoxides or in the case of hydroxychlorination of alkenes [Lopez et al. J. Am. Chem. Soc., 2017]. This project was supported by an ANR funding (CrystalBALL).

Now we would like to develop the concept of cascade catalysis in cristallo on oxidative transformations of alkenes, performed in multi-active sites cross-linked artificial enzyme crystals (CLEC). It will consist of an evolution of our mastered CLEC application that will require innovative chemical modifications on NikA protein crystals themselves for the insertion of a second inorganic catalytic site. Our knowledge in high performance of alkenes’ degradation using dioxygen as oxidant will drive us to add downstream transformations in an original cascade of catalytic oxidation reactions, implying carbonation, aldehyde oxidation or degradation of polyunsaturated biomolecules. The project is divided into two main objectives: first, the construction of two catalytic sites within a protein, distinguished by their mode of insertion. One will be inserted via supramolecular interactions as mastered by the consortium today, while the other will be covalently linked at the level of the solvent channels of the protein crystals. The second objective will be a dual transformation of alkenes, oxidative cleavage into aldehydes then oxidation to carboxylic acids or oxidation to epoxides then transformation into carbonates. The oxidation of polyenes are an illustration of this strategy. This project was supported by an ANR funding (Ni(k)AGARA).
The biohybrid complexity can also be enhanced by the addition of a photosensitizer to design solid photocatalysts based on protein scaffolds such as LEAFY.

DNA as chiral inducer
C. Marchi-Delapierre

We propose to develop novel and eco-friendly catalysts for stereoselective transformations based on G-quadruplex (G4) nucleic acids constrained in a single controlled topology (RG4) and metal complexes. It has been demonstrated that the G4 topology, known to be quite diverse, has a great influence on the obtained enantiomeric excess. However, these different topologies are in constant equilibrium in the experimental conditions (nature of the buffer, cation content, pH, etc.), making the rationalization of the G4 efficiency as pre-catalysts rather difficult, in terms of both efficiency per se and opportunities to improve the process (recyclability, enantioselectivity, etc.). Our main goal consists of chemically constrain G4s into a single topology (RG4) to prevent structural interconversions. Two strategies will be implemented herein to keep on expanding the scope of RG4-based catalysis. First, Our home made synthetic metals-complexes, known and used for developing artificial metalloenzymes, will be used with RG4 pre-catalysts; to date, only a few metal complexes (mostly copper(II) based) has been used for DNAzyme-type reactions. Second, we will develop brand new enantioselective DNAzyme-type reactions, thanks to the addition of chiral cofactors, here TASQ (for template-assembled synthetic G-quartets). TASQ have indeed been demonstrated to be valuable DNAzyme boosting agents, sandwiching the cofactor (i.e., metal complexes) in between RG4 and TASQ, therefore creating a binding pocket optimized for catalysis. Using chiral TASQ will thus afford the possibility of catalyzing chiral reactions of outmost interest for industry-driven applications. (ANR CoolCAT, collaboration with N. Spinelli, DCM, UGA Grenoble and D. Monchaud, ICMUB, UB Dijon).