Engineered bidirectional promoters enable rapid multi-gene co-expression optimization

Author:

Vogl ThomasORCID,Kickenweiz Thomas,Pitzer Julia,Sturmberger Lukas,Weninger Astrid,Biggs Bradley W.ORCID,Köhler Eva-Maria,Baumschlager ArminORCID,Fischer Jasmin Elgin,Hyden Patrick,Wagner Marlies,Baumann Martina,Borth Nicole,Geier Martina,Ajikumar Parayil Kumaran,Glieder Anton

Abstract

AbstractNumerous synthetic biology endeavors require well-tuned co-expression of functional components for success. Classically, monodirectional promoters (MDPs) have been used for such applications, but MDPs are limited in terms of multi-gene co-expression capabilities. Consequently, there is a pressing need for new tools with improved flexibility in terms of genetic circuit design, metabolic pathway assembly, and optimization. Here, motivated by nature’s use of bidirectional promoters (BDPs) as a solution for efficient gene co-expression, we generate a library of 168 synthetic BDPs in the yeast Komagataella phaffii (syn. Pichia pastoris), leveraging naturally occurring BDPs as a parts repository. This library of synthetic BDPs allows for rapid screening of diverse expression profiles and ratios to optimize gene co-expression, including for metabolic pathways (taxadiene, β-carotene). The modular design strategies applied for creating the BDP library could be relevant in other eukaryotic hosts, enabling a myriad of metabolic engineering and synthetic biology applications.

Publisher

Springer Science and Business Media LLC

Subject

General Physics and Astronomy,General Biochemistry, Genetics and Molecular Biology,General Chemistry

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