This title appears in the Scientific Report : 2014 

Engineering of Corynebacterium glutamicum towards utilization of Methanol as carbon and energy source
Witthoff, Sabrina (Corresponding Author)
Biotechnologie; IBG-1
2015
Jülich Forschungszentrum Jülich GmbH Zentralbibliothek, Verlag 2015
V, 113 pp
Universität Düsseldorf, Diss., 2014
978-3-95806-029-6
Book
Dissertation / PhD Thesis
Addenda
Schriften des Forschungszentrums Jülich Reihe Gesundheit / Health 78
OpenAccess
OpenAccess
Please use the identifier: http://hdl.handle.net/2128/8532 in citations.
Methanol is a pure and inexpensive raw material, which is mainly produced from fossil-fuelbasedsynthesis gas. Over the past years, new approaches were developed for its productionfrom renewable carbon sources. In the chemical industry, methanol is already an importantcarbon feedstock, but it has found only limited application in biotechnology. This canpredominantly be attributed to the inability of important microbial platform organisms to utilizethis C1 compound. With the aim to make methanol a suitable substrate for microbial productionprocesses, the non-methylotrophic and industrially important amino acid-producing bacteriumCorynebacterium glutamicum was engineered towards the utilization of methanol as auxiliarycarbon source in a sugar-based medium.Initial experiments on the response of C. glutamicum to methanol showed that this organismis able to oxidize methanol to CO2 during the stationary phase with a rate of 0.83 ± 0.2 mM/h(2.8 ± 0.5 nmol min-1 mg CDW-1) in glucose/methanol defined medium. Methanol oxidation wasshown to be subject to carbon catabolite repression in the presence of glucose and to bedependent on the transcriptional regulator RamA. Global gene expression studies revealed thatthe alcohol dehydrogenase gene adhA as well as the aldehyde dehydrogenase gene ald were upregulatedin the presence of methanol. Analysis of a mutant lacking the adhA gene showed a67% reduced methanol consumption rate (0.27 ± 0.05 mM/h), indicating that AdhA is mainlyresponsible for the oxidation of methanol to formaldehyde. The oxidation of formaldehyde toformate was found to be catalyzed predominantly by two enzymes, the acetaldehydedehydrogenase Ald and the mycothiol-dependent formaldehyde dehydrogenase AdhE. Adouble mutant lacking ald and adhE was severely impaired in its ability to oxidizeformaldehyde. The oxidation of formate to CO2 is catalyzed by formate dehydrogenase (FDH).Deletion of fdhF (annotated as FDH) and fdhD (annotated as FDH accessory protein) inC. glutamicum abolished formate oxidation and resulted in an increased formate sensitivity.Growth studies with molybdenum and tungsten indicated that FdhF is a molybdenumdependentenzyme. The electron acceptor of FdhF is not NAD(P)+ and still unknown. ...