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Increasing lysergic acid levels for ergot alkaloid biosynthesis: Directing catalysis via the F-G loop of Clavine oxidases

Most ergot alkaloid drugs are semi-synthetically derived from the natural product lysergic acid, a valuable precursor for the development of novel ergot alkaloid drugs. Clavine oxidase (CloA) is a putative cytochrome P450, identified in the ergot alkaloid biosynthesis pathway, and a key enzyme that...

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Autores principales: Lim, Li Rong, Wong, Garrett, Go, Maybelle K., Yew, Wen Shan
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10060963/
https://www.ncbi.nlm.nih.gov/pubmed/37007471
http://dx.doi.org/10.3389/fmicb.2023.1150937
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author Lim, Li Rong
Wong, Garrett
Go, Maybelle K.
Yew, Wen Shan
author_facet Lim, Li Rong
Wong, Garrett
Go, Maybelle K.
Yew, Wen Shan
author_sort Lim, Li Rong
collection PubMed
description Most ergot alkaloid drugs are semi-synthetically derived from the natural product lysergic acid, a valuable precursor for the development of novel ergot alkaloid drugs. Clavine oxidase (CloA) is a putative cytochrome P450, identified in the ergot alkaloid biosynthesis pathway, and a key enzyme that catalyzes the formation of lysergic acid from the precursor alkaloid agroclavine in a two-step oxidation reaction. We demonstrated in this study that Saccharomyces cerevisiae can be used as a viable host for the functional expression of CloA from Claviceps purpurea and its orthologs. We also showed that CloA orthologs differ in their ability to oxidize the substrate agroclavine, with some orthologs only able to perform the first oxidation reaction to produce elymoclavine. Of particular note, we identified a region between the F-G helices of the enzyme that may be involved in directing oxidation of agroclavine by substrate recognition and uptake. Using this knowledge, engineered CloAs were shown to produce lysergic acid at levels exceeding that of wildtype CloA orthologs; a CloA variant, chimeric AT5 9Hypo CloA, increased production levels of lysergic acid to 15 times higher as compared to the wildtype enzyme, demonstrating future utility for the industrial production of ergot alkaloids using biosynthetic routes.
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spelling pubmed-100609632023-03-31 Increasing lysergic acid levels for ergot alkaloid biosynthesis: Directing catalysis via the F-G loop of Clavine oxidases Lim, Li Rong Wong, Garrett Go, Maybelle K. Yew, Wen Shan Front Microbiol Microbiology Most ergot alkaloid drugs are semi-synthetically derived from the natural product lysergic acid, a valuable precursor for the development of novel ergot alkaloid drugs. Clavine oxidase (CloA) is a putative cytochrome P450, identified in the ergot alkaloid biosynthesis pathway, and a key enzyme that catalyzes the formation of lysergic acid from the precursor alkaloid agroclavine in a two-step oxidation reaction. We demonstrated in this study that Saccharomyces cerevisiae can be used as a viable host for the functional expression of CloA from Claviceps purpurea and its orthologs. We also showed that CloA orthologs differ in their ability to oxidize the substrate agroclavine, with some orthologs only able to perform the first oxidation reaction to produce elymoclavine. Of particular note, we identified a region between the F-G helices of the enzyme that may be involved in directing oxidation of agroclavine by substrate recognition and uptake. Using this knowledge, engineered CloAs were shown to produce lysergic acid at levels exceeding that of wildtype CloA orthologs; a CloA variant, chimeric AT5 9Hypo CloA, increased production levels of lysergic acid to 15 times higher as compared to the wildtype enzyme, demonstrating future utility for the industrial production of ergot alkaloids using biosynthetic routes. Frontiers Media S.A. 2023-03-16 /pmc/articles/PMC10060963/ /pubmed/37007471 http://dx.doi.org/10.3389/fmicb.2023.1150937 Text en Copyright © 2023 Lim, Wong, Go and Yew. 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 Microbiology
Lim, Li Rong
Wong, Garrett
Go, Maybelle K.
Yew, Wen Shan
Increasing lysergic acid levels for ergot alkaloid biosynthesis: Directing catalysis via the F-G loop of Clavine oxidases
title Increasing lysergic acid levels for ergot alkaloid biosynthesis: Directing catalysis via the F-G loop of Clavine oxidases
title_full Increasing lysergic acid levels for ergot alkaloid biosynthesis: Directing catalysis via the F-G loop of Clavine oxidases
title_fullStr Increasing lysergic acid levels for ergot alkaloid biosynthesis: Directing catalysis via the F-G loop of Clavine oxidases
title_full_unstemmed Increasing lysergic acid levels for ergot alkaloid biosynthesis: Directing catalysis via the F-G loop of Clavine oxidases
title_short Increasing lysergic acid levels for ergot alkaloid biosynthesis: Directing catalysis via the F-G loop of Clavine oxidases
title_sort increasing lysergic acid levels for ergot alkaloid biosynthesis: directing catalysis via the f-g loop of clavine oxidases
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10060963/
https://www.ncbi.nlm.nih.gov/pubmed/37007471
http://dx.doi.org/10.3389/fmicb.2023.1150937
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