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

Biotechnology and Bioengineering, vol. 121, pp. 1789–1802

Abstract

Abstract 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.

Authors 9

1. Alvaro R. Lara corresponding

   Aarhus University

   Affiliation as printed

   Department of Biological and Chemical Engineering Aarhus University Aarhus Denmark

   Department of Biological and Chemical Engineering, Aarhus University, Aarhus, Denmark

2. Flavio Kunert Aachen Biochemical Engineering (AVT.BioVT)

   RWTH Aachen University

   Affiliation as printed

   Biochemical Engineering (AVT.BioVT) RWTH Aachen University Aachen Germany

   Biochemical Engineering (AVT.BioVT), RWTH Aachen University, Aachen, Germany

3. Vincent Vandenbroucke

   University of Liège · Gembloux Agro-Bio Tech

   Affiliation as printed

   Terra Research and Teaching Centre, Microbial Processes and Interactions (MiPI), Gembloux Agro‐Bio Tech University of Liège Gembloux Belgium

4. Hilal Taymaz‐Nikerel

   Istanbul Bilgi University

   Affiliation as printed

   Department of Genetics and Bioengineering Istanbul Bilgi University Istanbul Turkey

   Department of Genetics and Bioengineering, Istanbul Bilgi University, Istanbul, Turkey

5. Luz María Martínez

   Universidad Nacional Autónoma de México

   Affiliation as printed

   Instituto de Biotecnología Universidad Nacional Autónoma de México Cuernavaca México

6. Juan‐Carlos Sigala

   Universidad Autónoma Metropolitana

   Affiliation as printed

   Departamento de Procesos y Tecnología Universidad Autónoma Metropolitana Ciudad de México México

7. Frank Delvigne

   University of Liège · Gembloux Agro-Bio Tech

   Affiliation as printed

   Terra Research and Teaching Centre, Microbial Processes and Interactions (MiPI), Gembloux Agro‐Bio Tech University of Liège Gembloux Belgium

8. Guillermo Gosset

   Universidad Nacional Autónoma de México

   Affiliation as printed

   Instituto de Biotecnología Universidad Nacional Autónoma de México Cuernavaca México

9. Jochen Büchs Aachen Biochemical Engineering (AVT.BioVT)

   RWTH Aachen University

   Affiliation as printed

   Biochemical Engineering (AVT.BioVT) RWTH Aachen University Aachen Germany

   Biochemical Engineering (AVT.BioVT), RWTH Aachen University, Aachen, Germany