Redesign allosterischer Regulation von Enzymen für eine selbst-regulierende und dynamische Kontrolle von Stoffflüssen in der mikrobiellen Produktion von Aminosäuren

The goal of this project is to redesign allosteric regulation of enzyme(s) for self-regulated and dynamic metabolic control in bioproduction processes. Using homoserine dehydrogenase (HSD) and lysine biosynthesis as a model system, we will implement lysine as a signalling molecule to control metabolic flux to the threonine pathway which is necessary for cell growth but undesired for lysine production. Different HSD variants with modified allosteric regulation will be designed. By construction of Corynebacterium glutamicum mutants bearing the mutated HSD, we will use metabolic and flux analyses to investigate the flux redistribution upon the genetic perturbation. With our approach, the enzyme activity of HSD should be adjusted according to the cellular physiological conditions during the bioprocess, especially to lysine concentration. At the stage of cell growth, the intracellular lysine concentration is low, and thus the inhibition of HSD will not be strong, allowing enough flux to the threonine pathways for cell growth. With the increase of lysine concentration in the production phase, the inhibition of HSD will be automatically enhanced and thus the substrate will be channelled into the pathway of lysine production. With this novel approach, we will develop a new tool for overcoming some of the major problems associated with drastic genetic modifications like gene overexpression or knockout in conventional strain development.

Stichworte

• Aminosäure
• Fermentationstechnik
• Protein engineering
• synthetische Biologie
• Systembiologie

Publikationen

• Bommareddy, Rajesh Reddy and Chen, Zhen and Rappert, Sugima and Zeng, An-Ping: A de novo NADPH generation pathway for improving lysine production of Corynebacterium glutamicum by rational design of the coenzyme specificity of glyceraldehyde 3-phosphate dehydrogenase. Metabolic engineering, 25: S. 30--7, sep 2014.
• Chen, Z and Meyer, W and Rappert, S and Sun, J and Zeng, A P: Coevolutionary analysis enabled rational deregulation of allosteric enzyme inhibition in Corynebacterium glutamicum for lysine production Appl Environ Microbiol, 77(13): S. 4352--4360, 2011.
• Chen, Z and Bommareddy, R R and Frank, D and Rappert, S and Zeng, A P: Deregulation of feedback inhibition of phosphoenolpyruvate carboxylase for improved lysine production in Corynebacterium glutamicum Appl Environ Microbiol, 80(4): S. 1388--1393, 2014.
• Ma, C W and Xiu, Z L and Zeng, A P: Discovery of intramolecular signal transduction network based on a new protein dynamics model of energy dissipation PLoS One, 7(2): S. e31529, 2012.
• Ma, C W and Xiu, Z L and Zeng, A P: Exploring signal transduction in heteromultimeric protein based on energy dissipation model J Biomol Struct Dyn, 2013.
• Chen, Z and Rappert, S and Sun, J and Zeng, A P: Integrating molecular dynamics and co-evolutionary analysis for reliable target prediction and deregulation of the allosteric inhibition of aspartokinase for amino acid production J Biotechnol, 154(4): S. 248--254, 2011.
• Chen, Z and Zeng, A P: Protein design in systems metabolic engineering for industrial strain development Biotechnol J, 8(5): S. 523--533, 2013.
• Chen, Zhen and Jandt, Uwe and Rappert, Sugima and Zeng, An-Ping: Proteindesign für die Entwicklung von industriellen Mikroorganismen BIOspektrum, 19(1): S. 99--101, feb 2013.
• Chen, Z and Rappert, S and Zeng, A P: Rational Design of Allosteric Regulation of Homoserine Dehydrogenase by a Non-Natural Inhibitor L-Lysine ACS Synth Biol, 2013.