Investigation of fatty aldehyde and alcohol synthesis from fatty acids by αDox- or CAR-expressing Escherichia coli

Maurer, Susanne; Schewe, Hendrik; Schrader, Jens; Buchhaupt, Markus

doi:10.1016/j.jbiotec.2019.08.011

Cited by 11 publications

(39 citation statements)

References 22 publications

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“…Meanwhile, neither C n-1 alcohols nor C n-1 FAs, produced by over-reduction or re-oxidation of fatty aldehydes by endogenous fatty aldehyde reductases or oxidases, respectively, were detected in this study although they were b Reactions were performed with 2-mM myristic acid with Triton X-100 (1%, v/v), initiated by purified enzymes (0.1 or 1 μg) at 25 °C under various pH values: 0.1-M sodium acetate (pH 4 to 6), 0.1-M sodium phosphate (pH 6 to 7), 0.1-M Tris-HCl (pH 7 to 9), and 0.1-M glycine-NaOH (pH 9 to 10). Activity was determined by the oxygen depletion assay at the initial time phase often produced by whole cells containing fatty aldehyde-producing α-DOX or CAR in previous studies (Foo et al 2017;Maurer et al 2019). The catalytic performance of CalDOX was found to be superior to that of LepDOX in the wholecell E. coli system (Fig.…”

Section: Whole-cell Biotransformations For the Synthesis Of Fatty Aldehydesmentioning

confidence: 87%

“…To date, most of the biosynthetic pathways for fatty aldehydes -either as the final product or as precursors to industrially relevant compounds -have been devised by introducing fatty acid acyl-CoA/ACP reductases (FARs) (Foo et al 2020;Lehtinen et al 2018Lehtinen et al , 2017Reiser and Somerville 1997) or carboxylic acid reductases (CARs) (Akhtar et al 2013;Maurer et al 2019;Wu et al 2021). Many attempts were made for the de novo synthesis of fatty aldehydes by introducing FAR in various organisms such as Escherichia coli, Saccharomyces cerevisiae, or Acinetobacter baylyi (Foo et al 2020;Lehtinen et al 2018Lehtinen et al , 2017Reiser and Somerville 1997).…”

Section: Introductionmentioning

confidence: 99%

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Two novel cyanobacterial α-dioxygenases for the biosynthesis of fatty aldehydes

Kim

Brack

Bayer

et al. 2021

Appl Microbiol Biotechnol

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Abstractα-Dioxygenases (α-DOXs) are known as plant enzymes involved in the α-oxidation of fatty acids through which fatty aldehydes, with a high commercial value as flavor and fragrance compounds, are synthesized as products. Currently, little is known about α-DOXs from non-plant organisms. The phylogenic analysis reported here identified a substantial number of α-DOX enzymes across various taxa. Here, we report the functional characterization and Escherichia coli whole-cell application of two novel α-DOXs identified from cyanobacteria: CalDOX from Calothrix parietina and LepDOX from Leptolyngbya sp. The catalytic behavior of the recombinantly expressed CalDOX and LepDOX revealed that they are heme-dependent like plant α-DOXs but exhibit activities toward medium carbon fatty acids ranging from C10 to C14 unlike plant α-DOXs. The in-depth molecular investigation of cyanobacterial α-DOXs and their application in an E. coli whole system employed in this study is useful not only for the understanding of the molecular function of α-DOXs, but also for their industrial utilization in fatty aldehyde biosynthesis.Key points• Two novel α-dioxygenases from Cyanobacteria are reported• Both enzymes prefer medium-chain fatty acids• Both enzymes are useful for fatty aldehyde biosynthesis Graphical abstract

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Section: Whole-cell Biotransformations For the Synthesis Of Fatty Aldehydesmentioning

confidence: 87%

Section: Introductionmentioning

confidence: 99%

Two novel cyanobacterial α-dioxygenases for the biosynthesis of fatty aldehydes

Kim

Brack

Bayer

et al. 2021

Appl Microbiol Biotechnol

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“…[5a,b] Thep resent study not only proposes the key role of the active site water derived from H 2 O 2 activation in the catalysis of CYP152 peroxygenases for the first time,but also provides an ew,s imple,a nd more productive enzymatic system for aldehyde biosynthesis compared with the reported biocatalytic aldehyde-producing systems. [21] An important question to be answered is whether the aldehyde-forming mechanism is also applicable for other a,b-unsaturated fatty acids with different chain length. To our delight, both P450 SPa and P450 BSb indeed produced the aldehyde products lauraldehyde (C 12 )a nd tridecanal (C 13 )a st he main products from trans-2tridecenoic acid (C 13 )a nd trans-2-tetradecenoic acid (C 14 ), respectively (Figures S124,S125).…”

Section: Discussionmentioning

confidence: 99%

Unexpected Reactions of α,β‐Unsaturated Fatty Acids Provide Insight into the Mechanisms of CYP152 Peroxygenases

Jiang

Peng

et al. 2021

Angewandte Chemie

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CYP152 peroxygenases catalyzed ecarboxylation and hydroxylation of fatty acids using H 2 O 2 as cofactor.T o understand the molecular basis for the chemo-and regioselectivity of these unique P450 enzymes,weanalyzethe activities of three CYP152 peroxygenases (OleT JE ,P 450 SPa ,P 450 BSb ) towards cis-and trans-dodecenoic acids as substrate probes. The unexpected 6S-hydroxylation of the trans-isomer and 4Rhydroxylation of the cis-isomer by OleT JE ,a nd molecular docking results suggest that the unprecedented selectivity is due to OleT JE spreference of C2ÀC3 cis-configuration. In addition to the common epoxideproducts,undecanal is the unexpected major product of P450 SPa and P450 BSb regardless of the cis/ trans-configuration of substrates.The combined H 2 18 O 2 tracing experiments,M Ds imulations,a nd QM/MM calculations unravel an unusual mechanism for Compound I-mediated aldehyde formation in whichthe active site water derived from H 2 O 2 activation is involved in the generation of af ourmembered ring lactone intermediate.T hese findings provide new insights into the unusual mechanisms of CYP152 peroxygenases.

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“…The explanation for decreased product titer might be increased toxicity of the shorter carbon‐chain compound compared to 1,12‐dodecanediol . Further, a study targeting C10‐C12 aldehydes could not limit overreduction to less than 80 % in E. coli MG00 of E. coli K‐12 MG1655 origin …”

Section: Introductionmentioning

confidence: 99%

“…[28] Further, a study targeting C10-C12 aldehydes could not limit overreduction to less than 80 % in E. coli MG00 of E. coli K-12 MG1655 origin. [29] Targeting aldehydes, Hansen et al established an enzyme cascade to produce vanillin glucoside which is the naturally occurring compound in vanilla pods, via de novo biosynthesis. [18] Additionally, yeast strains used for vanillin-β-d-glucoside production were engineered to minimize overreduction to vanillyl alcohol.…”

Section: Introductionmentioning

confidence: 99%

In Vivo Reduction of Medium‐ to Long‐Chain Fatty Acids by Carboxylic Acid Reductase (CAR) Enzymes: Limitations and Solutions

Horvat

Winkler

2020

ChemCatChem

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Fatty aldehyde production by chemical synthesis causes an immense burden to the environment. Within this study, we explored a sustainable, aldehyde-selective and mild alternative approach by utilizing carboxylic acid reductases (CARs). CARs from Neurospora crassa (NcCAR), Thermothelomyces thermophila (TtCAR), Nocardia iowensis (NiCAR), Mycobacterium marinum (MmCAR) and Trametes versicolor (TvCAR) were overexpressed in E. coli K-12 MG1655 RARE (DE3) and screened for medium-to long-chain fatty acid (C6-C18) reduction. MmCAR showed the broadest tolerance towards all carbon-chain lengths and was selected for further investigations of fatty aldehyde synthesis in whole cells. To yield relevant product concentrations, different limitations of CAR whole-cell conversions were elucidated and compensated. We coupled an in vitro cofactor recycling system to a whole-cell biocatalyst to support cofactor supply and achieved 12.36 g L À 1 of octanal (STY 0.458 g L À 1 h À 1) with less than 1.5 % of 1-octanol.

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Investigation of fatty aldehyde and alcohol synthesis from fatty acids by αDox- or CAR-expressing Escherichia coli

Cited by 11 publications

References 22 publications

Two novel cyanobacterial α-dioxygenases for the biosynthesis of fatty aldehydes

Two novel cyanobacterial α-dioxygenases for the biosynthesis of fatty aldehydes

Unexpected Reactions of α,β‐Unsaturated Fatty Acids Provide Insight into the Mechanisms of CYP152 Peroxygenases

In Vivo Reduction of Medium‐ to Long‐Chain Fatty Acids by Carboxylic Acid Reductase (CAR) Enzymes: Limitations and Solutions

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