Proteome reallocation enables the selective de novo biosynthesis of non-linear, branched-chain acetate esters

Hyeongmin Seo; Richard J. Giannone; Yung-Hun Yang; Cong T. Trinh

doi:10.1016/j.ymben.2022.05.003

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Proteome reallocation enables the selective de novo biosynthesis of non-linear, branched-chain acetate esters

Journal article

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Proteome reallocation enables the selective de novo biosynthesis of non-linear, branched-chain acetate esters

Hyeongmin Seo, Richard J. Giannone, Yung-Hun Yang and Cong T. Trinh

Metabolic engineering, Vol.73, pp.38-49

09/01/2022

DOI: 10.1016/j.ymben.2022.05.003

PMID: 35561848

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Abstract

The one-carbon recursive ketoacid elongation pathway is responsible for making various branched-chain amino acids, aldehydes, alcohols, ketoacids, and acetate esters in living cells. Controlling selective microbial biosynthesis of these target molecules at high efficiency is challenging due to enzyme promiscuity, regulation, and metabolic burden. In this study, we present a systematic modular design approach to control proteome reallocation for selective microbial biosynthesis of branched-chain acetate esters. Through pathway modularization, we partitioned the branched-chain ester pathways into four submodules including ketoisovalerate submodule for converting pyruvate to ketoisovalerate, ketoacid elongation submodule for producing longer carbon-chain ketoacids, ketoacid decarboxylase submodule for converting ketoacids to alcohols, and alcohol acyltransferase submodule for producing branched-chain acetate esters by condensing alcohols and acetyl-CoA. By systematic manipulation of pathway gene replication and transcription, enzyme specificity of the first committed steps of these submodules, and downstream competing pathways, we demonstrated selective microbial production of isoamyl acetate over isobutyl acetate. We found that the optimized isoamyl acetate pathway globally redistributed the amino acid fractions in the proteomes and required up to 23-31% proteome reallocation at the expense of other cellular resources, such as those required to generate precursor metabolites and energy for growth and amino acid biosynthesis. From glucose fed-batch fermentation, the engineered strains produced isoamyl acetate up to a titer of 8.8 g/L (>0.25 g/L toxicity limit), a yield of 0.22 g/g (61% of maximal theoretical value), and 86% selectivity, achieving the highest titers, yields and selectivity of isoamyl acetate reported to date.

Biotechnology & Applied Microbiology

Life Sciences & Biomedicine

Science & Technology

Details

Title: Subtitle: Proteome reallocation enables the selective de novo biosynthesis of non-linear, branched-chain acetate esters
Creators: Hyeongmin Seo - Oak Ridge National Laboratory
Richard J. Giannone - Oak Ridge National Laboratory
Yung-Hun Yang - Konkuk University
Cong T. Trinh - University of Tennessee at Knoxville
Resource Type: Journal article
Publication Details: Metabolic engineering, Vol.73, pp.38-49
DOI: 10.1016/j.ymben.2022.05.003
PMID: 35561848
NLM abbreviation: Metab Eng
ISSN: 1096-7176
eISSN: 1096-7184
Publisher: Elsevier
Number of pages: 12
Grant note: DE-SC0022226 / DOE BER award; United States Department of Energy (DOE) DE-AC02-05CH11231 / U.S. Department of Energy Joint Genome Institute, a DOE Office of Science User Facility; United States Department of Energy (DOE) U.S. Department of Energy Bioenergy Research Center - Office of Biological and Environmental Research in the DOE Office of Science; United States Department of Energy (DOE) DE-AC05-000R22725 / DOE subcontract grant by the Center of Bioenergy Innovation; United States Department of Energy (DOE)
Language: English
Date published: 09/01/2022
Academic Unit: Chemical and Biochemical Engineering
Record Identifier: 9984696582402771

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