Transport-controlled growth decoupling for self-induced protein expression with a glycerol-repressible genetic circuit.

Autor: Lara AR; Department of Biological and Chemical Engineering, Aarhus University, Aarhus, Denmark., Kunert F; Biochemical Engineering (AVT.BioVT), RWTH Aachen University, Aachen, Germany., Vandenbroucke V; Terra Research and Teaching Centre, Microbial Processes and Interactions (MiPI), Gembloux Agro-Bio Tech, University of Liège, Gembloux, Belgium., Taymaz-Nikerel H; Department of Genetics and Bioengineering, Istanbul Bilgi University, Istanbul, Turkey., Martínez LM; Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, México., Sigala JC; Departamento de Procesos y Tecnología, Universidad Autónoma Metropolitana, Ciudad de México, México., Delvigne F; Terra Research and Teaching Centre, Microbial Processes and Interactions (MiPI), Gembloux Agro-Bio Tech, University of Liège, Gembloux, Belgium., Gosset G; Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, México., Büchs J; Biochemical Engineering (AVT.BioVT), RWTH Aachen University, Aachen, Germany.
Jazyk: angličtina
Zdroj: Biotechnology and bioengineering [Biotechnol Bioeng] 2024 Jun; Vol. 121 (6), pp. 1789-1802. Date of Electronic Publication: 2024 Mar 12.
DOI: 10.1002/bit.28697
Abstrakt: Decoupling cell formation from recombinant protein synthesis is a potent strategy to intensify bioprocesses. Escherichia coli strains with mutations in the glucose uptake components lack catabolite repression, display low growth rate, no overflow metabolism, and high recombinant protein yields. Fast growth rates were promoted by the simultaneous consumption of glucose and glycerol, and this was followed by a phase of slow growth, when only glucose remained in the medium. A glycerol-repressible genetic circuit was designed to autonomously induce recombinant protein expression. The engineered strain bearing the genetic circuit was cultured in 3.9 g L -1 glycerol + 18 g L -1 glucose in microbioreactors with online oxygen transfer rate monitoring. The growth was fast during the simultaneous consumption of both carbon sources (C-sources), while expression of the recombinant protein was low. When glycerol was depleted, the growth rate decreased, and the specific fluorescence reached values 17% higher than those obtained with a strong constitutive promoter. Despite the relatively high amount of C-source used, no oxygen limitation was observed. The proposed approach eliminates the need for the substrate feeding or inducers addition and is set as a simple batch culture while mimicking fed-batch performance.
(© 2024 The Authors. Biotechnology and Bioengineering published by Wiley Periodicals LLC.)
Databáze: MEDLINE
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