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Multidrug resistance gene regulators as scaffolds for the design and evolution of artificial metalloenzymes

Objective

Enzymes are remarkable catalysts. The unmatched rate accelerations and exacting selectivities that these protein molecules achieve have whetted the appetite of chemists to harness the prowess of enzyme catalysis for industrial applications. However, natural enzymes can only catalyze a small fraction of the reactions routinely employed by synthetic chemists. As a result, creating designer biocatalysts with the ability to efficiently catalyze transformations not found in nature’s repertoire is a long-standing goal in chemical biology. To meet this challenge, this proposal describes our plans to generate proficient enzymes for palladium-catalyzed cross-coupling reactions. Specifically, we will create hybrid catalysts by recruiting active palladium complexes to the promiscuous binding sites of multidrug resistance gene regulators. We will validate productive assemblies by a rigorous biophysical characterization and evaluate the resulting artificial metalloenzymes for their ability to catalyze model Suzuki-Miyaura cross-coupling reactions. To refine the activities and selectivities of these primitive catalysts we will explore directed evolution protocols to identify mutations in the protein scaffolds that are beneficial for catalysis. In one strategy, we will establish a fluorescence-based screening approach that allows for testing libraries of hybrid catalysts in multi-well format. Another strategy will evaluate the possibility of performing cross-coupling reaction in vivo. Toward this end, artificial metalloenzymes will be assembled in the periplasm and utilized for the synthesis of a non-standard amino acid, which subsequently can be incorporated into a selection marker. As a result, bacteria producing improved variants will outgrow those with less efficient catalysts under selection conditions. Overall, our efforts will generate proficient designer enzymes that could prove valuable for applications in sustainable chemical processes.

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Keywords

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Programme(s)

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Topic(s)

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Funding Scheme

Funding scheme (or “Type of Action”) inside a programme with common features. It specifies: the scope of what is funded; the reimbursement rate; specific evaluation criteria to qualify for funding; and the use of simplified forms of costs like lump sums.

MSCA-IF - Marie Skłodowska-Curie Individual Fellowships (IF)

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Call for proposal

Procedure for inviting applicants to submit project proposals, with the aim of receiving EU funding.

(opens in new window) H2020-MSCA-IF-2016

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Coordinator

RIJKSUNIVERSITEIT GRONINGEN
Net EU contribution

Net EU financial contribution. The sum of money that the participant receives, deducted by the EU contribution to its linked third party. It considers the distribution of the EU financial contribution between direct beneficiaries of the project and other types of participants, like third-party participants.

€ 165 598,80
Address
Broerstraat 5
9712CP Groningen
Netherlands

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Activity type
Higher or Secondary Education Establishments
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Total cost

The total costs incurred by this organisation to participate in the project, including direct and indirect costs. This amount is a subset of the overall project budget.

€ 165 598,80
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