Integration of proteomics and metabolomics into the Design, Build, Test, Learn cycle to improve 3-hydroxypropionic acid production in Aspergillus pseudoterreus

Pomraning, Kyle R.; Dai, Ziyu; Munoz Munoz, Nathalie; Kim, Young-Mo; Gao, Yuqian; Deng, Shuang; Kim, Joonhoon; Hofstad, Beth A.; Swita, Marie S.; Lemmon, Teresa L.; Collett, James R.; Panisko, Ellen A.; Webb-Robertson, Bobbie-Jo M.; Zucker, Jeremy D.; Nicora, Carrie D.; De Paoli, Henrique; Baker, Scott E.; Burnum-Johnson, Kristin E.; Hillson, Nathan J.; Magnuson, Jon K.

doi:10.3389/fbioe.2021.603832

Integration of proteomics and metabolomics into the Design, Build, Test, Learn cycle to improve 3-hydroxypropionic acid production in Aspergillus pseudoterreus

Journal Article · Wed Apr 07 00:00:00 EDT 2021 · Frontiers in Bioengineering and Biotechnology

DOI:https://doi.org/10.3389/fbioe.2021.603832· OSTI ID:1779764

^[1]; Dai, Ziyu ^[1]; Munoz Munoz, Nathalie ^[1]; ^[1]; Gao, Yuqian ^[1]; Deng, Shuang ^[1]; ^[1]; Hofstad, Beth A. ^[1]; Swita, Marie S. ^[1]; Lemmon, Teresa L. ^[1]; Collett, James R. ^[1]; Panisko, Ellen A. ^[2]; Webb-Robertson, Bobbie-Jo M. ^[1]; ^[1]; ^[1]; De Paoli, Henrique ^[3]; ^[1]; ^[1]; Hillson, Nathan J. ^[4]; ^[1]

BATTELLE (PACIFIC NW LAB)
HANFORD LABORATORY MGMT & INTEGRATION
Lawrence Berkeley National Laboratory
Lawrence Berkeley National Lab

Biological engineering of microorganisms to produce value-added chemicals is a promising route to sustainable manufacturing. However, overproduction of metabolic intermediates at high titer, rate, and yield from inexpensive substrates is challenging in non-model systems where limited information is available regarding metabolic flux and its control in production conditions. Integrated multi-omic analyses of engineered strains offers an in-depth look at metabolites and proteins directly involved in growth and production of target and non-target bioproducts. Here we applied multi-omic analyses to overproduction of the polymer precursor 3-hydroxypropionic acid (3HP) in the filamentous fungus Aspergillus pseudoterreus. A synthetic pathway consisting of aspartate decarboxylase, beta-alanine pyruvate transaminase, and 3HP dehydrogenase was designed and built for A. pseudoterreus. Strains with single- and multi-copy integration events were isolated and multi-omics analysis consisting of intracellular and extracellular metabolomics and targeted and global proteomics was used to interrogate the strains in shake-flask and bioreactor conditions. Production of a variety of co-products (organic acids and glycerol) and oxidative degradation of 3HP were identified as metabolic pathways competing with 3HP production. Intracellular accumulation of nitrogen as 2,4-diaminobutanoate was identified as an off-target nitrogen sink that may also limit flux through the engineered 3HP pathway. Elimination of the high-expression oxidative 3HP degradation pathway by deletion of a putative malonate semialdehyde dehydrogenase improved the yield of 3HP by 3.4× after 10 days in shake-flask culture. This is the first report of 3HP production in a filamentous fungus amenable to industrial scale biomanufacturing of organic acids at high titer and low pH.

Research Organization:: Pacific Northwest National Laboratory (PNNL), Richland, WA (United States)

Sponsoring Organization:: USDOE

DOE Contract Number:: AC05-76RL01830

OSTI ID:: 1779764

Report Number(s):: PNNL-SA-149911

Journal Information:: Frontiers in Bioengineering and Biotechnology, Journal Name: Frontiers in Bioengineering and Biotechnology Vol. 9

Country of Publication:: United States

Language:: English

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