Heterologous erythromycin production across strain and plasmid construction

Abstract

The establishment of erythromycin production within the heterologous host E. coli marked an accomplishment in genetic transfer capacity. Namely, over 20 genes and 50 kb of DNA was introduced to E. coli for successful heterologous biosynthetic reconstitution. However, the prospect for production levels that approach those of the native host requires the application of engineering tools associated with E. coli. In this report, metabolic and genomic engineering were implemented to improve the E. coli cellular background and the plasmid platform supporting heterologous erythromycin formation. Results include improved plasmid stability and metabolic support for biosynthetic product formation. Specifically, the new plasmid design for erythromycin formation allowed for ≥89% stability relative to current standards (20% stability). In addition, the new strain (termed LF01) designed to improve carbon flow to the erythromycin biosynthetic pathway provided a 400% improvement in titer level. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 34:271-276, 2018.

Marine Biosurfactants: Biosynthesis, Structural Diversity and Biotechnological Applications

“…However, the burden of plasmid replication can lead to certain instability of constructs [230,239,240]. Strategies have been developed to address this [241], but it is altogether abrogated by chromosomal integration strategies. Here, diverse tools are available like random chromosomal integration carried out using transposon Tn5 [242] or site-specific integration using transposon Tn7 or targeted recombination [238,243], with all methods being functional in a range of bacterial species.…”

Leveraging synthetic biology for producing bioactive polyketides and non-ribosomal peptides in bacterial heterologous hosts

“…143,144 The most recent study achieved an erythromycin titer of 40 mg L −1 . 145 A recently developed strategy for cloning BGCs into heterologous hosts resulted in the heterologous production of cyanobacterial non-ribosomal peptides in E. coli. 146 Greunke et al used Direct Pathway Cloning (DiPAC) to refactor the BGC for anabaenopeptin with E. coli-specific promoters and introduced the optimized pathway into E. coli via two replicative plasmids.…”

Acyltransferases as Tools for Polyketide Synthase Engineering

“…Finally, the efficient production of natural and engineered polyketides often requires the tuning of the native producer strain or the development of suitable heterologous expression hosts [ 182 , 239 , 240 ]. Preferably, diverse actinomycetes [ 241 , 242 , 243 , 244 , 245 ], E. coli [ 246 , 247 , 248 ], Bacillus [ 249 ], Saccharomyces [ 250 , 251 ] and/or Aspergillus strains [ 252 , 253 ] were used for heterologous expression of polyketide pathways in the past.…”