Combinatorial optimization of gene expression through recombinase-mediated promoter and terminator shuffling in yeast.
Charlotte CautereelsJolien SmetsPeter W BirchamDries De RuysscherAnna ZimmermannPeter De RijkKevin J VerstrepenAnton GorkovskiyJoleen MasscheleinKevin J VerstrepenPublished in: Nature communications (2024)
Microbes are increasingly employed as cell factories to produce biomolecules. This often involves the expression of complex heterologous biosynthesis pathways in host strains. Achieving maximal product yields and avoiding build-up of (toxic) intermediates requires balanced expression of every pathway gene. However, despite progress in metabolic modeling, the optimization of gene expression still heavily relies on trial-and-error. Here, we report an approach for in vivo, multiplexed Gene Expression Modification by LoxPsym-Cre Recombination (GEMbLeR). GEMbLeR exploits orthogonal LoxPsym sites to independently shuffle promoter and terminator modules at distinct genomic loci. This approach facilitates creation of large strain libraries, in which expression of every pathway gene ranges over 120-fold and each strain harbors a unique expression profile. When applied to the biosynthetic pathway of astaxanthin, an industrially relevant antioxidant, a single round of GEMbLeR improved pathway flux and doubled production titers. Together, this shows that GEMbLeR allows rapid and efficient gene expression optimization in heterologous biosynthetic pathways, offering possibilities for enhancing the performance of microbial cell factories.
Keyphrases
- gene expression
- dna methylation
- poor prognosis
- genome wide
- single cell
- copy number
- clinical trial
- binding protein
- escherichia coli
- dna damage
- transcription factor
- microbial community
- stem cells
- saccharomyces cerevisiae
- long non coding rna
- body composition
- phase iii
- resistance training
- network analysis
- phase ii
- cell wall