A tailor-made, self-sufficient and recyclable monooxygenase catalyst based on coimmobilized cytochrome P450 BM3 and glucose dehydrogenase

Donya Valikhani, Juan M. Bolivar, Alexander Dennig, Bernd Nidetzky

Publikation: Beitrag in einer FachzeitschriftArtikelForschungBegutachtung

Abstract

Cytochrome P450 monooxygenases (P450s) promote hydroxylations in a broad variety of substrates. Their prowess in C–H bond functionalization renders P450s promising catalysts for organic synthesis. However, operating P450 reactions involve complex management of the main substrates, O2 and nicotinamide adenine dinucleotide phosphate (NAD(P)H) reducing equivalents against an overall background of low operational stability. Whole-cell biocatalysis, although often used, offers no general solution to these problems. Herein, we present the design of a tailor-made, self-sufficient, operationally stabilized and recyclable P450 catalyst on porous solid support. Using enzymes as fusion proteins with the polycationic binding module Zbasic2, the P450s BM3 (from Bacillus megaterium) was coimmobilized with glucose dehydrogenase (type IV; from B. megaterium) on anionic sulfopropyl-activated carrier (ReliSorb SP). Immobilization via Zbasic2 enabled each enzyme to be loaded in controllable amount, thus maximizing the relative portion of the rate limiting P450 BM3 (up to 19.5 U/gcarrier) in total enzyme immobilized. Using lauric acid as a representative P450 substrate that is poorly accessible to whole-cell catalysts, we demonstrate complete hydroxylation at low catalyst loading (≤0.1 mol%) and efficient electron coupling (74%), inside of the catalyst particle, to the regeneration of NADPH from glucose (27 cycles) was achieved. The immobilized P450 BM3 showed a total turnover number of ∼18,000, thus allowing active catalyst to be recycled up to 20 times. This study therefore supports the idea of practical heterogeneous catalysis by P450s systems immobilized on solid support.

Spracheenglisch
Seiten2416-2425
Seitenumfang10
FachzeitschriftBiotechnology and Bioengineering
Jahrgang115
Ausgabennummer10
DOIs
StatusVeröffentlicht - 1 Okt 2018

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Glucose 1-Dehydrogenase
Mixed Function Oxygenases
Cytochrome P-450 Enzyme System
Glucose
Bacillus megaterium
Catalysts
lauric acid
Hydroxylation
NADP
Catalyst supports
Enzymes
Biocatalysis
Synthetic Chemistry Techniques
Immobilized Enzymes
Substrates
Catalysis
Immobilization
Regeneration
Bacilli
Electrons

Schlagwörter

    ASJC Scopus subject areas

    • Biotechnology
    • Bioengineering
    • !!Applied Microbiology and Biotechnology

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    A tailor-made, self-sufficient and recyclable monooxygenase catalyst based on coimmobilized cytochrome P450 BM3 and glucose dehydrogenase. / Valikhani, Donya; Bolivar, Juan M.; Dennig, Alexander; Nidetzky, Bernd.

    in: Biotechnology and Bioengineering, Jahrgang 115, Nr. 10, 01.10.2018, S. 2416-2425.

    Publikation: Beitrag in einer FachzeitschriftArtikelForschungBegutachtung

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    AU - Valikhani, Donya

    AU - Bolivar, Juan M.

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    AU - Nidetzky, Bernd

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