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Genetic engineering of an industrial yeast Candida glycerinogenes for efficient production of 2-phenylethanol.

Abstract：

:Microbial cell factories offer an economic approach for synthesizing "natural'" aromatic flavor compounds. During their fermentation process, the inefficient synthesis pathway and product cytotoxicity are the major barriers to the high-level production. This study combined metabolic engineering and tolerance engineering strategies to maximize the valuable rose-smell 2-phenylethanol (2-PE) production in Candida glycerinogenes, a GRAS diploid industrial yeast. Firstly, 2-PE metabolic networks involved in Ehrlich pathway were stepwise rewired using metabolic engineering, including the following: (1) overexpressing L-phenylalanine permease Aap9 enhanced precursor uptake; (2) overexpressing enzymes (aminotransferase Aro9 and decarboxylase Aro10) of Ehrlich pathway increased catalytic efficiency; and (3) disrupting the formation of by-product phenylacetate catalyzed by Ald2 and Ald3 maximized the metabolic flux toward 2-PE. Then, tolerance engineering was applied by overexpression of a stress-inducible gene SLC1 in the metabolically engineered strain to further enhance 2-PE production. Combining these two approaches finally resulted in 5.0 g/L 2-PE in shake flasks, with productivity reaching 0.21 g/L/h, which were increased by 38.9% and 177% compared with those of the non-engineered strain, respectively. The 2-PE yield of this engineered strain was 0.71 g/g L-phenylalanine, corresponding to 95.9% of theoretical yield. This study provides a reference to efficiently engineering of microbial cell factories for other valuable aromatic compounds. KEY POINTS: • Metabolic engineering improved 2-PE biosynthesis. • Tolerance engineering alleviated product inhibition, contributing to 2-PE production. • The best strain produced 5.0 g/L 2-PE with 0.959 mol/mol yield and high productivity.

journal_name

Appl Microbiol Biotechnol

journal_title

Applied microbiology and biotechnology

authors

Wang Y,Zhang Z,Lu X,Zong H,Zhuge B

doi

10.1007/s00253-020-10991-4

subject

Has Abstract

pub_date

2020-12-01 00:00:00

pages

10481-10491

issue

eissn

0175-7598

issn

1432-0614

pii

10.1007/s00253-020-10991-4

journal_volume

104

pub_type

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Genetic engineering of an industrial yeast Candida glycerinogenes for efficient production of 2-phenylethanol.