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Vitamin B6 Production in Bacillus subtilis

Interference of Heterologous and Host Pathways

by Jonathan Rosenberg
Doctoral thesis
Date of Examination:2018-01-11
Date of issue:2018-01-23
Advisor:Dr. Fabian Moritz Commichau
Referee:Prof. Dr. Jörg Stülke
Referee:Prof. Dr. Carsten Lüder
crossref-logoPersistent Address: http://dx.doi.org/10.53846/goediss-6683

 

 

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Abstract

English

Fermentative production of valuable goods is an economically competitive and ecologically sustainable alternative to chemical synthesis. Several previous studies have been dedicated to the establishment of vitamin B6 production strains, but none was successful to compete with chemical synthesis. Fermentative production of vitamin B6 is hampered by toxic intermediates, a toxic end-product and insufficient knowledge about vitamin B6 homeostasis and related proteins and pathways. This work is dedicated to the identification of deleterious interactions between the heterologous production pathway and the host metabolism, and proposes and applies an approach to design novel, hybrid production pathways combining host underground metabolism and partial heterologous pathways. Mutant derivatives of the Gram-positive model bacterium Bacillus subtilis, which are able to grow in presence of toxic amounts of pyridoxal 5’- phosphate are shown to carry mutations leading to deregulation of the biotin biosynthesis operon. Furthermore, the inhibitor of the threonine synthesis operon ThrR, which is also involved in resistance to the vitamin B6 synthesis intermediate 4-hydroxy-threonine phosphate, is characterized. In order to optimize an existing B6 production strain, targeted evolution is applied leading to a novel, non-native biosynthesis pathway of endogenous underground and heterologous enzyme activities. This approach is regarded to be ubiquitously applicable in different species and for other products and the characterization of the host-heterologous interferences is of great value for future projects for fermentative B6 production.
Keywords: Vitamin B6; Fermentation; Production of valuable goods; Pyridoxal; Pyridoxine; Underground Metabolism; Metabolic Pathways; Suppressor Mutants
 

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