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Anaerobic Fungal Mevalonate Pathway Genomic Biases Lead to Heterologous Toxicity Underpredicted by Codon Adaptation Indices

Anaerobic fungi are emerging biotechnology platforms with genomes rich in biosynthetic potential. Yet, the heterologous expression of their biosynthetic pathways has had limited success in model hosts like E. coli. We find one reason for this is that the genome composition of anaerobic fungi like P....

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Autores principales: Hillman, Ethan T., Frazier, Elizabeth M., Shank, Evan K., Ortiz-Velez, Adrian N., Englaender, Jacob A., Solomon, Kevin V.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8468974/
https://www.ncbi.nlm.nih.gov/pubmed/34576881
http://dx.doi.org/10.3390/microorganisms9091986
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author Hillman, Ethan T.
Frazier, Elizabeth M.
Shank, Evan K.
Ortiz-Velez, Adrian N.
Englaender, Jacob A.
Solomon, Kevin V.
author_facet Hillman, Ethan T.
Frazier, Elizabeth M.
Shank, Evan K.
Ortiz-Velez, Adrian N.
Englaender, Jacob A.
Solomon, Kevin V.
author_sort Hillman, Ethan T.
collection PubMed
description Anaerobic fungi are emerging biotechnology platforms with genomes rich in biosynthetic potential. Yet, the heterologous expression of their biosynthetic pathways has had limited success in model hosts like E. coli. We find one reason for this is that the genome composition of anaerobic fungi like P. indianae are extremely AT-biased with a particular preference for rare and semi-rare AT-rich tRNAs in E coli, which are not explicitly predicted by standard codon adaptation indices (CAI). Native P. indianae genes with these extreme biases create drastic growth defects in E. coli (up to 69% reduction in growth), which is not seen in genes from other organisms with similar CAIs. However, codon optimization rescues growth, allowing for gene evaluation. In this manner, we demonstrate that anaerobic fungal homologs such as PI.atoB are more active than S. cerevisiae homologs in a hybrid pathway, increasing the production of mevalonate up to 2.5 g/L (more than two-fold) and reducing waste carbon to acetate by ~90% under the conditions tested. This work demonstrates the bioproduction potential of anaerobic fungal enzyme homologs and how the analysis of codon utilization enables the study of otherwise difficult to express genes that have applications in biocatalysis and natural product discovery.
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spelling pubmed-84689742021-09-27 Anaerobic Fungal Mevalonate Pathway Genomic Biases Lead to Heterologous Toxicity Underpredicted by Codon Adaptation Indices Hillman, Ethan T. Frazier, Elizabeth M. Shank, Evan K. Ortiz-Velez, Adrian N. Englaender, Jacob A. Solomon, Kevin V. Microorganisms Article Anaerobic fungi are emerging biotechnology platforms with genomes rich in biosynthetic potential. Yet, the heterologous expression of their biosynthetic pathways has had limited success in model hosts like E. coli. We find one reason for this is that the genome composition of anaerobic fungi like P. indianae are extremely AT-biased with a particular preference for rare and semi-rare AT-rich tRNAs in E coli, which are not explicitly predicted by standard codon adaptation indices (CAI). Native P. indianae genes with these extreme biases create drastic growth defects in E. coli (up to 69% reduction in growth), which is not seen in genes from other organisms with similar CAIs. However, codon optimization rescues growth, allowing for gene evaluation. In this manner, we demonstrate that anaerobic fungal homologs such as PI.atoB are more active than S. cerevisiae homologs in a hybrid pathway, increasing the production of mevalonate up to 2.5 g/L (more than two-fold) and reducing waste carbon to acetate by ~90% under the conditions tested. This work demonstrates the bioproduction potential of anaerobic fungal enzyme homologs and how the analysis of codon utilization enables the study of otherwise difficult to express genes that have applications in biocatalysis and natural product discovery. MDPI 2021-09-18 /pmc/articles/PMC8468974/ /pubmed/34576881 http://dx.doi.org/10.3390/microorganisms9091986 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Hillman, Ethan T.
Frazier, Elizabeth M.
Shank, Evan K.
Ortiz-Velez, Adrian N.
Englaender, Jacob A.
Solomon, Kevin V.
Anaerobic Fungal Mevalonate Pathway Genomic Biases Lead to Heterologous Toxicity Underpredicted by Codon Adaptation Indices
title Anaerobic Fungal Mevalonate Pathway Genomic Biases Lead to Heterologous Toxicity Underpredicted by Codon Adaptation Indices
title_full Anaerobic Fungal Mevalonate Pathway Genomic Biases Lead to Heterologous Toxicity Underpredicted by Codon Adaptation Indices
title_fullStr Anaerobic Fungal Mevalonate Pathway Genomic Biases Lead to Heterologous Toxicity Underpredicted by Codon Adaptation Indices
title_full_unstemmed Anaerobic Fungal Mevalonate Pathway Genomic Biases Lead to Heterologous Toxicity Underpredicted by Codon Adaptation Indices
title_short Anaerobic Fungal Mevalonate Pathway Genomic Biases Lead to Heterologous Toxicity Underpredicted by Codon Adaptation Indices
title_sort anaerobic fungal mevalonate pathway genomic biases lead to heterologous toxicity underpredicted by codon adaptation indices
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8468974/
https://www.ncbi.nlm.nih.gov/pubmed/34576881
http://dx.doi.org/10.3390/microorganisms9091986
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