Avermectin B1a production in Streptomyces avermitilis is enhanced by engineering aveC and precursor supply genes

Avermectins (AVEs) are economically potent anthelmintic agents produced by Streptomyces avermitilis . Among eight AVE components, B1a exhibits the highest insecticidal activity. The purpose of this study was to enhance B1a production, particularly in the high-yielding industrial strain A229, by a co...

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Bibliographic Details
Published in:Applied microbiology and biotechnology 2022-03, Vol.106 (5-6), p.2191-2205
Main Authors: Hao, Yi, You, Yanting, Chen, Zhi, Li, Jilun, Liu, Gang, Wen, Ying
Format: Article
Language:English
Subjects:
Amino acids
Analysis
Anthelmintic agents
Anthelmintics
Antiparasitic agents
Applied Microbial and Cell Physiology
Avermectin
Biomedical and Life Sciences
Biotechnology
Carbon dioxide
Carboxylation
Chemical synthesis
Composition
FADD protein
Gene expression
Genes
Genetic aspects
Identification and classification
Industrial strains
Life Sciences
Methods
Methylmalonyl-CoA
Microbial Genetics and Genomics
Microbiology
Mutation
Oxidation
Precursors
Properties
Streptomyces
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Summary:Avermectins (AVEs) are economically potent anthelmintic agents produced by Streptomyces avermitilis . Among eight AVE components, B1a exhibits the highest insecticidal activity. The purpose of this study was to enhance B1a production, particularly in the high-yielding industrial strain A229, by a combination strategy involving the following steps. (i) aveC gene was engineered to increase B1a:B2a ratio. Three aveC variants ( aveC2m , aveC5m , and aveC8m , respectively encoding two, five, and eight amino acid mutations) were synthesized by fusion PCR. B1a:B2a ratio in A229 derivative having kasOp*- controlled aveC8m reached 1.33 (B1a and B2a titers were 8120 and 6124 μg/mL). Corresponding values in A229 were 0.99 and 6447 and 6480 μg/mL. (ii) β -oxidation pathway genes fadD and fadAB were overexpressed in wild-type (WT) strain and A229 to increase supply of acyl-CoA precursors for AVE production. The resulting strains all showed increased B1a titer. Co-overexpression of pkn5p -driven fadD and fadAB in A229 led to B1a titer of 8537 μg/mL. (iii) Genes bicA and ecaA involved in cyanobacterial CO 2 -concentrating mechanism (CCM) were introduced into WT and A229 to enhance carboxylation velocity of acetyl-CoA and propionyl-CoA carboxylases, leading to increased supply of malonyl- and methylmalonyl-CoA precursors and increased B1a titer. Co-expression of bicA and ecaA in A229 led to B1a titer of 8083 μg/mL. (iv) aveC8m , fadD - fadAB , and bicA - ecaA were co-overexpressed in A229, resulting in maximal B1a titer (9613 μg/mL; 49.1% increase relative to A229). Our findings demonstrate that the combination strategy we provided here is an efficient approach for improving B1a production in industrial strains. Key points • aveC mutation increased avermectin B1a:B2a ratio and B1a titer. • Higher levels of acyl-CoA precursors contributed to enhanced B1a production. • B1a titer in an industrial strain was increased by 49.1% via a combination strategy.
ISSN:0175-7598
1432-0614
DOI:10.1007/s00253-022-11854-w