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Heterogeneous catalysis with artificial metalloenzyme crystals

Researchers at the Chemistry and Biology of Metals laboratory are working on artificial metalloenzymes, one of which is a hydride composed of the NikA protein and complexes of iron or ruthenium. In order to improve their properties, they managed to stabilize the crystals of NikA by crosslinking. This resulting solid hybrid catalyst makes it possible to stabilize the artificial catalytic site within the protein crystal.

Published on 20 February 2018
An artificial metalloenzyme (ArM) is a hybrid construct resulting from the insertion of an inorganic synthetic complex within a DNA or protein biomolecule. The whole forms an eco-compatible catalyst that works in mild conditions in accordance with the principles of green chemistry. However, an additional step can be taken by proposing these catalysts in a heterogeneous (liquid-solid) version.

The Bioinspired Chemistry and Environment team [Chemistry and Biology of Metals laboratory] designs ArMs in order to propose new sustainable catalysis methodologies for oxidation reactions. In this context, this team has developed several powerful systems based in particular on the NikA protein (a nickel transport protein without enzymatic properties) and complexes of iron and ruthenium. However, ArMs can be further improved in order to widen the substrate range (limited by aqueous phase solubility), the number of catalytic cycles (limited by the stability of the proteins in an oxidizing medium) and the diversity of reactions (willingness to access combinations of catalytic processes).
To achieve these goals, the researchers of this team chose to create an original heterogeneous version of ArMs. This work could be achieved thanks to the stabilization of NikA protein crystals by cross-linking (Cross-Linked Enzyme Crystals: CLEC). A first step consisted in the development of cross-linking conditions of ArMs’ crystals suitable for a substantial production of catalytic crystals (ArM-CLEC) related to the use of 50 to 100 crystals per experiment. Then, the ArM-CLEC being stable in water-organic solvent mixtures (from 4 to 70°C and in the presence of numerous oxidants), their catalytic properties could be tested. An efficiency comparable to that of a "natural" enzyme has been demonstrated in particular with regard to the oxidative cleavage reaction of styrene derivatives (

An ArM-CLEC crystal consists of NikA-FeL2 hybrids (FeL2 being an iron complex with a phenolate ligand that confers the purple color to the crystals) cross-linked with glutaraldehyde. The stability of the crystals in a mixture of water-acetonitrile-1-1 solvent makes it possible to use many lipophilic substrates and to test various oxidation reactions, here oxidative cleavage and hydroxychlorination.

Thus, the corresponding aldehyde could be obtained with more than 28,000 TON (TurnOver Number) thanks to 50 recycling steps carried out with the same batch of crystals, guarantying the stability of the heterogeneous material. By way of comparison, the hybrid under homogeneous catalysis conditions only led to 140 TON. The great originality of this solid hybrid catalyst is thus to allow the stabilization of the artificial catalytic site within the protein crystal in addition to the discovering of a new reactivity for this organic complex and artificial enzymes in general.
Researchers now aim to provide even more eco-friendly catalytic systems that combine different catalysts. This study illustrates the power of the association of homogeneous, heterogeneous and biological catalysts.

This work brings together synthesis, catalysis and crystallography of proteins studies. It results from a collaboration between two teams from the Chemistry and Biology of Metals laboratory: the Bioinspired Chemistry and Environment team and the BioCatalysis team.
This work is funded by the ANR CrystalBall and an ARCANE Labex granted thesis.

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