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Title: Accelerating pathway evolution by increasing the gene dosage of chromosomal segments

Abstract

Experimental evolution is a critical tool in many disciplines, including metabolic engineering and synthetic biology. However, current methods rely on the chance occurrence of a key step that can dramatically accelerate evolution in natural systems, namely increased gene dosage. Our studies sought to induce the targeted amplification of chromosomal segments to facilitate rapid evolution. Since increased gene dosage confers novel phenotypes and genetic redundancy, we developed a method, Evolution by Amplification and Synthetic Biology (EASy), to create tandem arrays of chromosomal regions. In Acinetobacter baylyi, EASy was demonstrated on an important bioenergy problem, the catabolism of lignin-derived aromatic compounds. The initial focus on guaiacol (2-methoxyphenol), a common lignin degradation product, led to the discovery of Amycolatopsis genes (gcoAB) encoding a cytochrome P450 enzyme that converts guaiacol to catechol. However, chromosomal integration of gcoAB in Pseudomonas putida or A. baylyi did not enable guaiacol to be used as the sole carbon source despite catechol being a growth substrate. In ~1,000 generations, EASy yielded alleles that in single chromosomal copy confer growth on guaiacol. Different variants emerged, including fusions between GcoA and CatA (catechol 1,2-dioxygenase). This study illustrates the power of harnessing chromosomal gene amplification to accelerate the evolution of desirable traits.

Authors:
; ; ; ; ; ; ; ; ; ; ORCiD logo; ORCiD logo
Publication Date:
Research Org.:
National Renewable Energy Lab. (NREL), Golden, CO (United States)
Sponsoring Org.:
USDOE Office of Energy Efficiency and Renewable Energy (EERE), Bioenergy Technologies Office (EE-3B)
OSTI Identifier:
1466555
Report Number(s):
NREL/JA-5100-71408
Journal ID: ISSN 0027-8424
DOE Contract Number:  
AC36-08GO28308
Resource Type:
Journal Article
Journal Name:
Proceedings of the National Academy of Sciences of the United States of America
Additional Journal Information:
Journal Volume: 115; Journal Issue: 27; Journal ID: ISSN 0027-8424
Publisher:
National Academy of Sciences, Washington, DC (United States)
Country of Publication:
United States
Language:
English
Subject:
09 BIOMASS FUELS; gene amplification; evolution; P450; guaiacol; Acinetobacter

Citation Formats

Tumen-Velasquez, Melissa, Johnson, Christopher W., Ahmed, Alaa, Dominick, Graham, Fulk, Emily M., Khanna, Payal, Lee, Sarah A., Schmidt, Alicia L., Linger, Jeffrey G., Eiteman, Mark A., Beckham, Gregg T., and Neidle, Ellen L. Accelerating pathway evolution by increasing the gene dosage of chromosomal segments. United States: N. p., 2018. Web. doi:10.1073/pnas.1803745115.
Tumen-Velasquez, Melissa, Johnson, Christopher W., Ahmed, Alaa, Dominick, Graham, Fulk, Emily M., Khanna, Payal, Lee, Sarah A., Schmidt, Alicia L., Linger, Jeffrey G., Eiteman, Mark A., Beckham, Gregg T., & Neidle, Ellen L. Accelerating pathway evolution by increasing the gene dosage of chromosomal segments. United States. doi:10.1073/pnas.1803745115.
Tumen-Velasquez, Melissa, Johnson, Christopher W., Ahmed, Alaa, Dominick, Graham, Fulk, Emily M., Khanna, Payal, Lee, Sarah A., Schmidt, Alicia L., Linger, Jeffrey G., Eiteman, Mark A., Beckham, Gregg T., and Neidle, Ellen L. Mon . "Accelerating pathway evolution by increasing the gene dosage of chromosomal segments". United States. doi:10.1073/pnas.1803745115.
@article{osti_1466555,
title = {Accelerating pathway evolution by increasing the gene dosage of chromosomal segments},
author = {Tumen-Velasquez, Melissa and Johnson, Christopher W. and Ahmed, Alaa and Dominick, Graham and Fulk, Emily M. and Khanna, Payal and Lee, Sarah A. and Schmidt, Alicia L. and Linger, Jeffrey G. and Eiteman, Mark A. and Beckham, Gregg T. and Neidle, Ellen L.},
abstractNote = {Experimental evolution is a critical tool in many disciplines, including metabolic engineering and synthetic biology. However, current methods rely on the chance occurrence of a key step that can dramatically accelerate evolution in natural systems, namely increased gene dosage. Our studies sought to induce the targeted amplification of chromosomal segments to facilitate rapid evolution. Since increased gene dosage confers novel phenotypes and genetic redundancy, we developed a method, Evolution by Amplification and Synthetic Biology (EASy), to create tandem arrays of chromosomal regions. In Acinetobacter baylyi, EASy was demonstrated on an important bioenergy problem, the catabolism of lignin-derived aromatic compounds. The initial focus on guaiacol (2-methoxyphenol), a common lignin degradation product, led to the discovery of Amycolatopsis genes (gcoAB) encoding a cytochrome P450 enzyme that converts guaiacol to catechol. However, chromosomal integration of gcoAB in Pseudomonas putida or A. baylyi did not enable guaiacol to be used as the sole carbon source despite catechol being a growth substrate. In ~1,000 generations, EASy yielded alleles that in single chromosomal copy confer growth on guaiacol. Different variants emerged, including fusions between GcoA and CatA (catechol 1,2-dioxygenase). This study illustrates the power of harnessing chromosomal gene amplification to accelerate the evolution of desirable traits.},
doi = {10.1073/pnas.1803745115},
journal = {Proceedings of the National Academy of Sciences of the United States of America},
issn = {0027-8424},
number = 27,
volume = 115,
place = {United States},
year = {2018},
month = {6}
}

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