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Terpene Hydroxylation with Microbial Cytochrome P450 Monooxygenases

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Book cover Biotechnology of Isoprenoids

Part of the book series: Advances in Biochemical Engineering/Biotechnology ((ABE,volume 148))

Abstract

Terpenoids comprise a highly diverse group of natural products. In addition to their basic carbon skeleton, they differ from one another in their functional groups. Functional groups attached to the carbon skeleton are the basis of the terpenoids’ diverse properties. Further modifications of terpene olefins include the introduction of acyl-, aryl-, or sugar moieties and usually start with oxidations catalyzed by cytochrome P450 monooxygenases (P450s, CYPs). P450s are ubiquitously distributed throughout nature, involved in essential biological pathways such as terpenoid biosynthesis as well as the tailoring of terpenoids and other natural products. Their ability to introduce oxygen into nonactivated C–H bonds is unique and makes P450s very attractive for applications in biotechnology. Especially in the field of terpene oxidation, biotransformation methods emerge as an attractive alternative to classical chemical synthesis. For this reason, microbial P450s depict a highly interesting target for protein engineering approaches in order to increase selectivity and activity, respectively. Microbial P450s have been described to convert industrial and pharmaceutically interesting terpenoids such as ionones, limone, valencene, resin acids, and triterpenes (including steroids) as well as vitamin D3. Highly selective and active mutants have been evolved by applying classical site-directed mutagenesis as well as directed evolution of proteins. As P450s usually depend on electron transfer proteins, mutagenesis has also been applied to improve the interactions between P450s and their respective redox partners. This chapter provides an overview of terpenoid hydroxylation reactions catalyzed by bacterial P450s and highlights the achievements made by protein engineering to establish productive hydroxylation processes.

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Notes

  1. 1.

    The difference between terpenes and terpenoids is that compounds belonging to the latter category contain additional functional groups whereas terpenes are solely composed of carbon and hydrogen. Cytochromes P450 are often involved in the further functionalization of terpenes into terpenoids, but terpenoids themselves can also be substrates for P450 enzymes. For this reason the terms terpenes and terpenoids are used synonymously throughout the chapter.

Abbreviations

Adx:

Adrenodoxin

P450s:

Cytochrome P450 enzymes

BM3:

Cytochrome P450 102A1

P450cam :

Cytochrome P450 101A1

CPR:

Cytochrome P450 reductase

DMSO:

Dimethylsulfoxide

epPCR:

Error-prone polymerase chain reaction

Fdx:

Ferredoxin

FdR:

Ferredoxin reductase

FMN:

Flavine mononucleotide

FAD:

Flavine adenine dinucleotide

NAD(P)H:

Nicotine amide adenine dinucleotide (phosphate)

Pdx:

Putidaredoxin

THF:

Tetrahydrofuran

VD3 :

Vitamin D3

1α,25(OH)2VD3 :

1α,25-dihydroxyvitamin D3

CYP:

cytochrome P450

KBA:

11-keto-β-boswellic acid

PDB:

Protein Data Bank

VDR:

Vitamin D receptor

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Acknowledgments:

The work is supported by a grant of the Deutsche Forschungsgemeinschaft to R.B. (Be 1343/23-1). S.J. is supported by the INTERREG program of the European Union, and D.S. by a grant from the Saarland Ministry of Economics and Science.

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  1. Department of Biochemistry, Saarland University, Campus B2 2, 66123, Saarbruecken, Germany

    Simon Janocha, Daniela Schmitz & Rita Bernhardt

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  1. Simon Janocha
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  2. Daniela Schmitz
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  3. Rita Bernhardt
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Correspondence to Rita Bernhardt .

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  1. Bioverfahrenstechnik, DECHEMA-Forschungsinstitut, Frankfurt am Main, Germany

    Jens Schrader

  2. Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia, Canada

    Jörg Bohlmann

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Janocha, S., Schmitz, D., Bernhardt, R. (2015). Terpene Hydroxylation with Microbial Cytochrome P450 Monooxygenases. In: Schrader, J., Bohlmann, J. (eds) Biotechnology of Isoprenoids. Advances in Biochemical Engineering/Biotechnology, vol 148. Springer, Cham. https://doi.org/10.1007/10_2014_296

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