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Engineering of CYP153A33 With Enhanced Ratio of Hydroxylation to Overoxidation Activity in Whole-Cell Biotransformation of Medium-Chain 1-Alkanols

α,ω-Dodecanediol is a versatile material that has been widely used not only as an adhesive and crosslinking reagent, but also as a building block in the pharmaceutical and polymer industries. The biosynthesis of α,ω-dodecanediol from fatty derivatives, such as dodecane and dodecanol, requires an ω-s...

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Autores principales: Park, Hyuna, Bak, Doyeong, Jeon, Wooyoung, Jang, Minjung, Ahn, Jung-Oh, Choi, Kwon-Young
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8764613/
https://www.ncbi.nlm.nih.gov/pubmed/35059390
http://dx.doi.org/10.3389/fbioe.2021.817455
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author Park, Hyuna
Bak, Doyeong
Jeon, Wooyoung
Jang, Minjung
Ahn, Jung-Oh
Choi, Kwon-Young
author_facet Park, Hyuna
Bak, Doyeong
Jeon, Wooyoung
Jang, Minjung
Ahn, Jung-Oh
Choi, Kwon-Young
author_sort Park, Hyuna
collection PubMed
description α,ω-Dodecanediol is a versatile material that has been widely used not only as an adhesive and crosslinking reagent, but also as a building block in the pharmaceutical and polymer industries. The biosynthesis of α,ω-dodecanediol from fatty derivatives, such as dodecane and dodecanol, requires an ω-specific hydroxylation step using monooxygenase enzymes. An issue with the whole-cell biotransformation of 1-dodecanol using cytochrome P450 monooxygenase (CYP) with ω-specific hydroxylation activity was the low conversion and production of the over-oxidized product of dodecanoic acid. In this study, CYP153A33 from Marinobacter aquaeolei was engineered to obtain higher ω-specific hydroxylation activity through site-directed mutagenesis. The target residue was mutated to increase flux toward α,ω-dodecanediol synthesis, while reducing the generation of the overoxidation product of dodecanoic acid and α,ω-dodecanedioic acid. Among the evaluated variants, CYP153A33 P136A showed a significant increase in 1-dodecanol conversion, i.e., 71.2% (7.12 mM from 10 mM 1-dodecanol), with an increased hydroxylation to over-oxidation activity ratio, i.e., 32.4. Finally, the applicability of this engineered enzyme for ω-specific hydroxylation against several 1-alkanols, i.e., from C6 to C16, was investigated and discussed based on the structure-activity relationship.
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spelling pubmed-87646132022-01-19 Engineering of CYP153A33 With Enhanced Ratio of Hydroxylation to Overoxidation Activity in Whole-Cell Biotransformation of Medium-Chain 1-Alkanols Park, Hyuna Bak, Doyeong Jeon, Wooyoung Jang, Minjung Ahn, Jung-Oh Choi, Kwon-Young Front Bioeng Biotechnol Bioengineering and Biotechnology α,ω-Dodecanediol is a versatile material that has been widely used not only as an adhesive and crosslinking reagent, but also as a building block in the pharmaceutical and polymer industries. The biosynthesis of α,ω-dodecanediol from fatty derivatives, such as dodecane and dodecanol, requires an ω-specific hydroxylation step using monooxygenase enzymes. An issue with the whole-cell biotransformation of 1-dodecanol using cytochrome P450 monooxygenase (CYP) with ω-specific hydroxylation activity was the low conversion and production of the over-oxidized product of dodecanoic acid. In this study, CYP153A33 from Marinobacter aquaeolei was engineered to obtain higher ω-specific hydroxylation activity through site-directed mutagenesis. The target residue was mutated to increase flux toward α,ω-dodecanediol synthesis, while reducing the generation of the overoxidation product of dodecanoic acid and α,ω-dodecanedioic acid. Among the evaluated variants, CYP153A33 P136A showed a significant increase in 1-dodecanol conversion, i.e., 71.2% (7.12 mM from 10 mM 1-dodecanol), with an increased hydroxylation to over-oxidation activity ratio, i.e., 32.4. Finally, the applicability of this engineered enzyme for ω-specific hydroxylation against several 1-alkanols, i.e., from C6 to C16, was investigated and discussed based on the structure-activity relationship. Frontiers Media S.A. 2022-01-03 /pmc/articles/PMC8764613/ /pubmed/35059390 http://dx.doi.org/10.3389/fbioe.2021.817455 Text en Copyright © 2022 Park, Bak, Jeon, Jang, Ahn and Choi. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Bioengineering and Biotechnology
Park, Hyuna
Bak, Doyeong
Jeon, Wooyoung
Jang, Minjung
Ahn, Jung-Oh
Choi, Kwon-Young
Engineering of CYP153A33 With Enhanced Ratio of Hydroxylation to Overoxidation Activity in Whole-Cell Biotransformation of Medium-Chain 1-Alkanols
title Engineering of CYP153A33 With Enhanced Ratio of Hydroxylation to Overoxidation Activity in Whole-Cell Biotransformation of Medium-Chain 1-Alkanols
title_full Engineering of CYP153A33 With Enhanced Ratio of Hydroxylation to Overoxidation Activity in Whole-Cell Biotransformation of Medium-Chain 1-Alkanols
title_fullStr Engineering of CYP153A33 With Enhanced Ratio of Hydroxylation to Overoxidation Activity in Whole-Cell Biotransformation of Medium-Chain 1-Alkanols
title_full_unstemmed Engineering of CYP153A33 With Enhanced Ratio of Hydroxylation to Overoxidation Activity in Whole-Cell Biotransformation of Medium-Chain 1-Alkanols
title_short Engineering of CYP153A33 With Enhanced Ratio of Hydroxylation to Overoxidation Activity in Whole-Cell Biotransformation of Medium-Chain 1-Alkanols
title_sort engineering of cyp153a33 with enhanced ratio of hydroxylation to overoxidation activity in whole-cell biotransformation of medium-chain 1-alkanols
topic Bioengineering and Biotechnology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8764613/
https://www.ncbi.nlm.nih.gov/pubmed/35059390
http://dx.doi.org/10.3389/fbioe.2021.817455
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