Using random chemical mutagenesis we obtained the mutant of Cupriavidus necator H16 which was capable of improved (about 35 %) production of poly(3-hydroxybuytrate) (PHB) compared to the wild-type strain. The mutant exhibited significantly enhanced specific activities of enzymes involved in oxidative stress response such as malic enzyme, NADP-dependent isocitrate dehydrogenase, glucose-6-phosphate dehydrogenase and glutamate dehydrogenase. Probably, due to the activation of these enzymes, we also observed an increase of NADPH/NADP⁺ ratio. It is likely that as a side effect of the increase of NADPH/NADP⁺ ratio the activity of PHB biosynthetic pathway was enhanced, which supported the accumulation of PHB. Furthermore, the mutant was also able to incorporate propionate into copolymer poly(3-hydroxybuytyrate-co-3-hydroxyvalerate) [P(3HB-co-3HV)] more efficiently than the wild-type strain (Y3HV/prec = 0.17 and 0.29 for the wild-type strain and the mutant, respectively)). We assume that it may be caused by lower availability of oxaloacetate for the utilization of propionyl-CoA in 2-methylcitrate cycle due to increased action of malic enzyme. Therefore, propionyl-CoA was incorporated into copolymer rather than transformed to pyruvate via 2-methylcitrate cycle. Thus, the mutant was capable of the utilization of waste frying oils and the production of P(3HB-co-3HV) with better yields and improved content of 3HV resulting in better mechanical properties of copolymer than the wild-type strain. The results of this work may be used for the development of innovative fermentation strategies for the production of PHA and also it might help to define novel targets for the genetic manipulations of PHA producing bacteria.
- MeSH
- acylkoenzym A metabolismus MeSH
- bakteriální proteiny genetika metabolismus MeSH
- bioreaktory MeSH
- biosyntetické dráhy MeSH
- Brassica rapa MeSH
- Cupriavidus necator účinky léků genetika metabolismus MeSH
- fermentace MeSH
- hydroxybutyráty metabolismus MeSH
- mutageneze * MeSH
- oleje rostlin chemie metabolismus MeSH
- oxidační stres MeSH
- polyestery metabolismus MeSH
- polyhydroxyalkanoáty metabolismus MeSH
- průmyslový odpad MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Waste rapeseed oil is a useful substrate for polyhydroxyalkanoates (PHA) production employing Cupriavidus necator H16. In fed-batch mode, we obtained biomass and PHA yields of 138 and 105 g l(-1), respectively. Yield coefficient and volumetric productivity were 0.83 g PHA per g oil and 1.46 g l(-1) h(-1), respectively. Propanol at 1% (v/v) enhanced both PHA and biomass formation significantly and, furthermore, resulted in incorporation of 3-hydroxyvalerate units into PHA structure. Thus, propanol can be used as an effective precursor of 3-hydroxyvalarete for production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer. During the fed-batch cultivation, propanol concentration was maintained at 1% which resulted in 8% content of 3-hydroxyvalerate in copolymer.
