Inside Cover: New Sesquiterpene Oxidations with CYP260A1 and CYP264B1 from <i>Sorangium cellulosum</i> So ce56 (ChemBioChem 18/2015)

Schifrin, Alexander; Litzenburger, Martin; Ringle, Michael; Ly, Thuy T. B.; Bernhardt, Rita

doi:10.1002/cbic.201500636

Cited by 3 publications

(4 citation statements)

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“…Among them, CYP260A1 is one of the novel P450s, which we have recently shown to perform a selective 1α‐hydroxylation of C‐19 steroids . In addition, the oxidation of sesquiterpenes and conversion of drugs by CYP260A1 was also demonstrated. Although its autologous redox partners Fdx2/FdRB and Fdx8/FdRB were identified as efficient for sesquiterpene oxidation , the bovine redox partners adrenodoxin (Adx 4–108 ) and adrenodoxin reductase (AdR) were shown to be efficient surrogate redox partners , and they were employed for the establishment of an Escherichia coli ‐based whole‐cell system .…”

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confidence: 99%

Structural characterization of CYP260A1 from Sorangium cellulosum to investigate the 1α‐hydroxylation of a mineralocorticoid

et al. 2016

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In this study, we report the crystal structure of the cytochrome P450 CYP260A1 (PDB 5LIV) from the myxobacterium Sorangium cellulosum So ce56. In addition, we investigated the hydroxylation of 11-deoxycorticosterone by CYP260A1 by reconstituting the enzyme with the surrogate redox partners adrenodoxin and adrenodoxin reductase. The major product of this steroid conversion was identified as 1a-hydroxy-11-deoxycorticosterone, a novel D4 C-21 steroidal derivative. Furthermore, we docked the substrate into the crystal structure and replaced Ser326, the residue responsible for substrate orientation, with asparagine and observed that the mutant S326N displayed higher activity and selectivity for the formation of 1a-hydroxy-11-deoxycorticosterone compared to the wild-type CYP260A1. Thus, our findings highlight the usefulness of the obtained crystal structure of CYP260A1 in identifying biotechnologically more efficient reactions.

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confidence: 99%

Structural characterization of CYP260A1 from Sorangium cellulosum to investigate the 1α‐hydroxylation of a mineralocorticoid

et al. 2016

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“…There are only few examples for biocatalysts with activity towards the tested sesquiterpenes such as α‐humulene, β‐bisabolene or premnaspirodiene, which makes the enzyme CYP109Q5 particularly interesting. The previous examples for the production of 5‐ or 8‐hydroxy‐α‐humulene were extended by the biocatalyst identified in this work (Yu et al ., ; Schifrin et al ., ). For the first time, oxyfunctionalization of β‐bisabolene was enabled by a heterologously expressed bacterial P450.…”

Section: Discussionmentioning

confidence: 97%

“…For the identification of novel terpene hydroxylases, we therefore searched for organisms that possess large numbers of potentially interesting P450s for the oxyfunctionalization of terpenes. Studies on the myxobacterium S. cellulosum So ce56 have already identified and characterized P450s, able to efficiently hydroxylate norisoprenoids and sesquiterpenes (Ly et al ., ; Schifrin et al ., ; Litzenburger and Bernhardt, ). Myxobacteria feature the largest bacterial genomes known today and a versatile secondary metabolism, making them particularly interesting for biotechnological applications (Schneiker et al ., ; Diez et al ., ).…”

Section: Discussionmentioning

confidence: 99%

Characterization and structure‐guided engineering of the novel versatile terpene monooxygenase CYP109Q5 from Chondromyces apiculatus DSM436

Klenk

Dubiel

Sharma

et al. 2018

Microbial Biotechnology

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Summary One of the major challenges in chemical synthesis is the selective oxyfunctionalization of non‐activated C‐H bonds, which can be enabled by biocatalysis using cytochrome P450 monooxygenases. In this study, we report on the characterization of the versatile CYP 109Q5 from Chondromyces apiculatus DSM 436, which is able to functionalize a wide range of substrates (terpenes, steroids and drugs), including the ring of β‐ionone in non‐allylic positions. The crystal structure of CYP 109Q5 revealed flexibility within the active site pocket that permitted the accommodation of bulky substrates, and enabled a structure‐guided approach to engineering the enzyme. Some variants of CYP 109Q5 displayed a switch in selectivity towards the non‐allylic positions of β‐ionone, allowing the simultaneous production of 2‐ and 3‐hydroxy‐β‐ionone, which are chemically challenging to synthesize and are important precursors for carotenoid synthesis. An efficient whole‐cell system finally enabled the production of up to 0.5 g l −1 hydroxylated products of β‐ionone; this system can be applied to product identification in further biotransformations. Overall, CYP 109Q5 proved to be highly evolvable and active. The studies in this work demonstrate that, using rational mutagenesis, the highly versatile CYP 109Q5 generalist can be progressively evolved to be an industrially valuable specialist for the synthesis of specific products.

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“…This enzyme was identified as a new class of bacterial P450 (Ewen et al 2009;Khatri et al 2016a) with a steroid hydroxylase activity towards C19 (testosterone and androstenedione) or C21 (11-deoxycorticosterone) type steroids at the very unique 1α-position in the presence of surrogate redox partners (adrenodoxin and adrenodoxin reductase) (Khatri et al 2016b;Ewen et al 2009;Schifrin et al 2015;Litzenburger et al 2015). However, the conversion of progesterone by CYP260A1 is very unselective (Khatri et al 2016a, b;Salamanca-Pinzon et al 2016), yielding a mixture of 1α-progesterone, 17α-progesterone and other products.…”

Section: Cyp260a1: Cytochrome P450 From Myxobacterium Sorangium Cellumentioning

confidence: 99%

Bacterial cytochrome P450-catalyzed regio- and stereoselective steroid hydroxylation enabled by directed evolution and rational design

Zhang

Peng

Zhao

et al. 2020

Bioresour. Bioprocess.

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Steroids are the most widely marketed products by the pharmaceutical industry after antibiotics. Steroid hydroxylation is one of the most important functionalizations because their derivatives enable a higher biological activity compared to their less polar non-hydroxylated analogs. Bacterial cytochrome P450s constitute promising biocatalysts for steroid hydroxylation due to their high expression level in common workhorses like Escherichia coli. However, they often suffer from wrong or insufficient regio-and/or stereoselectivity, low activity, narrow substrate range as well as insufficient thermostability, which hampers their industrial application. Fortunately, these problems can be generally solved by protein engineering based on directed evolution and rational design. In this work, an overview of recent developments on the engineering of bacterial cytochrome P450s for steroid hydroxylation is presented.

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Inside Cover: New Sesquiterpene Oxidations with CYP260A1 and CYP264B1 from Sorangium cellulosum So ce56 (ChemBioChem 18/2015)

Cited by 3 publications

References 0 publications

Structural characterization of CYP260A1 from Sorangium cellulosum to investigate the 1α‐hydroxylation of a mineralocorticoid

Structural characterization of CYP260A1 from Sorangium cellulosum to investigate the 1α‐hydroxylation of a mineralocorticoid

Characterization and structure‐guided engineering of the novel versatile terpene monooxygenase CYP109Q5 from Chondromyces apiculatus DSM436

Bacterial cytochrome P450-catalyzed regio- and stereoselective steroid hydroxylation enabled by directed evolution and rational design

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