Dietary Change Enables Robust Growth-Coupling of Heterologous Methyltransferase Activity in Yeast

Anne Sofie Lærke Hansen; Maitreya J. Dunham; Dushica Arsovska; Jie Zhang; Jay D. Keasling; Markus J. Herrgard; Michael K. Jensen

doi:10.1021/acssynbio.0c00348

Dietary Change Enables Robust Growth-Coupling of Heterologous Methyltransferase Activity in Yeast

Anne Sofie Lærke Hansen, Maitreya J. Dunham, Dushica Arsovska, Jie Zhang, Jay D. Keasling, Markus J. Herrgard, Michael K. Jensen^*

^*Corresponding author for this work

University of Washington

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Abstract

Genetic modifications of living organisms and proteins are made possible by a catalogue of molecular and synthetic biology tools, yet proper screening assays for genetic variants of interest continue to lag behind. Synthetic growth-coupling (GC) of enzyme activities offers a simple, inexpensive way to track such improvements. In this follow-up study we present the optimization of a recently established GC design for screening of heterologous methyltransferases (MTases) and related pathways in the yeast Saccharomyces cerevisiae. Specifically, upon testing different media compositions and genetic backgrounds, improved GC of different heterologous MTase activities is obtained. Furthermore, we demonstrate the strength of the system by screening a library of catechol O-MTase variants converting protocatechuic acid into vanillic acid. We demonstrated high correlation (R² = 0.775) between vanillic acid and cell density as a proxy for MTase activity. We envision that the improved MTase GC can aid evolution-guided optimization of biobased production processes for methylated compounds with yeast in the future.

Original language	English
Journal	ACS Synthetic Biology
Volume	9
Issue number	12
Pages (from-to)	3408-3415
Number of pages	8
ISSN	2161-5063
DOIs	https://doi.org/10.1021/acssynbio.0c00348
Publication status	Published - 2020

Keywords

Growth-coupling
Methyltransferase
Yeast
Synthetic biology
Natural products
Screening

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title = "Dietary Change Enables Robust Growth-Coupling of Heterologous Methyltransferase Activity in Yeast",

abstract = "Genetic modifications of living organisms and proteins are made possible by a catalogue of molecular and synthetic biology tools, yet proper screening assays for genetic variants of interest continue to lag behind. Synthetic growth-coupling (GC) of enzyme activities offers a simple, inexpensive way to track such improvements. In this follow-up study we present the optimization of a recently established GC design for screening of heterologous methyltransferases (MTases) and related pathways in the yeast Saccharomyces cerevisiae. Specifically, upon testing different media compositions and genetic backgrounds, improved GC of different heterologous MTase activities is obtained. Furthermore, we demonstrate the strength of the system by screening a library of catechol O-MTase variants converting protocatechuic acid into vanillic acid. We demonstrated high correlation (R2 = 0.775) between vanillic acid and cell density as a proxy for MTase activity. We envision that the improved MTase GC can aid evolution-guided optimization of biobased production processes for methylated compounds with yeast in the future.",

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T1 - Dietary Change Enables Robust Growth-Coupling of Heterologous Methyltransferase Activity in Yeast

AU - Hansen, Anne Sofie Lærke

AU - Dunham, Maitreya J.

AU - Arsovska, Dushica

AU - Zhang, Jie

AU - Keasling, Jay D.

AU - Herrgard, Markus J.

AU - Jensen, Michael K.

PY - 2020

Y1 - 2020

N2 - Genetic modifications of living organisms and proteins are made possible by a catalogue of molecular and synthetic biology tools, yet proper screening assays for genetic variants of interest continue to lag behind. Synthetic growth-coupling (GC) of enzyme activities offers a simple, inexpensive way to track such improvements. In this follow-up study we present the optimization of a recently established GC design for screening of heterologous methyltransferases (MTases) and related pathways in the yeast Saccharomyces cerevisiae. Specifically, upon testing different media compositions and genetic backgrounds, improved GC of different heterologous MTase activities is obtained. Furthermore, we demonstrate the strength of the system by screening a library of catechol O-MTase variants converting protocatechuic acid into vanillic acid. We demonstrated high correlation (R2 = 0.775) between vanillic acid and cell density as a proxy for MTase activity. We envision that the improved MTase GC can aid evolution-guided optimization of biobased production processes for methylated compounds with yeast in the future.

AB - Genetic modifications of living organisms and proteins are made possible by a catalogue of molecular and synthetic biology tools, yet proper screening assays for genetic variants of interest continue to lag behind. Synthetic growth-coupling (GC) of enzyme activities offers a simple, inexpensive way to track such improvements. In this follow-up study we present the optimization of a recently established GC design for screening of heterologous methyltransferases (MTases) and related pathways in the yeast Saccharomyces cerevisiae. Specifically, upon testing different media compositions and genetic backgrounds, improved GC of different heterologous MTase activities is obtained. Furthermore, we demonstrate the strength of the system by screening a library of catechol O-MTase variants converting protocatechuic acid into vanillic acid. We demonstrated high correlation (R2 = 0.775) between vanillic acid and cell density as a proxy for MTase activity. We envision that the improved MTase GC can aid evolution-guided optimization of biobased production processes for methylated compounds with yeast in the future.

KW - Growth-coupling

KW - Methyltransferase

KW - Yeast

KW - Synthetic biology

KW - Natural products

KW - Screening

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DO - 10.1021/acssynbio.0c00348

M3 - Journal article

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SN - 2161-5063

VL - 9

SP - 3408

EP - 3415

JO - ACS Synthetic Biology

JF - ACS Synthetic Biology

IS - 12

ER -

Dietary Change Enables Robust Growth-Coupling of Heterologous Methyltransferase Activity in Yeast

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