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Article

Adaptive Laboratory Evolution of Cupriavidus necator H16 for Carbon Co-Utilization with Glycerol

1
Department of Chemical & Biological Engineering and Advanced Biomanufacturing Centre, University of Sheffield, Sir Robert Hadfield Building, Mappin Street, Sheffield S1 3JD, UK
2
Croda Europe Ltd., Oak Road, Clough Road, Hull HU6 7PH, UK
3
National Center for Genetic Engineering and Biotechnology, 113 Thailand Science Park, Phahonyothin Road, Khlong Luang, Pathum Thani 12120, Thailand
*
Authors to whom correspondence should be addressed.
Int. J. Mol. Sci. 2019, 20(22), 5737; https://doi.org/10.3390/ijms20225737
Received: 26 September 2019 / Revised: 11 November 2019 / Accepted: 13 November 2019 / Published: 15 November 2019
Cupriavidus necator H16 is a non-pathogenic Gram-negative betaproteobacterium that can utilize a broad range of renewable heterotrophic resources to produce chemicals ranging from polyhydroxybutyrate (biopolymer) to alcohols, alkanes, and alkenes. However, C. necator H16 utilizes carbon sources to different efficiency, for example its growth in glycerol is 11.4 times slower than a favorable substrate like gluconate. This work used adaptive laboratory evolution to enhance the glycerol assimilation in C. necator H16 and identified a variant (v6C6) that can co-utilize gluconate and glycerol. The v6C6 variant has a specific growth rate in glycerol 9.5 times faster than the wild-type strain and grows faster in mixed gluconate–glycerol carbon sources compared to gluconate alone. It also accumulated more PHB when cultivated in glycerol medium compared to gluconate medium while the inverse is true for the wild-type strain. Through genome sequencing and expression studies, glycerol kinase was identified as the key enzyme for its improved glycerol utilization. The superior performance of v6C6 in assimilating pure glycerol was extended to crude glycerol (sweetwater) from an industrial fat splitting process. These results highlight the robustness of adaptive laboratory evolution for strain engineering and the versatility and potential of C. necator H16 for industrial waste glycerol valorization. View Full-Text
Keywords: Cupriavidus necator H16; Ralstonia eutropha H16; adaptive evolution; carbon co-utilization; glycerol; biodiesel; fat splitting; biopolymer Cupriavidus necator H16; Ralstonia eutropha H16; adaptive evolution; carbon co-utilization; glycerol; biodiesel; fat splitting; biopolymer
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MDPI and ACS Style

González-Villanueva, M.; Galaiya, H.; Staniland, P.; Staniland, J.; Savill, I.; Wong, T.S.; Tee, K.L. Adaptive Laboratory Evolution of Cupriavidus necator H16 for Carbon Co-Utilization with Glycerol. Int. J. Mol. Sci. 2019, 20, 5737. https://doi.org/10.3390/ijms20225737

AMA Style

González-Villanueva M, Galaiya H, Staniland P, Staniland J, Savill I, Wong TS, Tee KL. Adaptive Laboratory Evolution of Cupriavidus necator H16 for Carbon Co-Utilization with Glycerol. International Journal of Molecular Sciences. 2019; 20(22):5737. https://doi.org/10.3390/ijms20225737

Chicago/Turabian Style

González-Villanueva, Miriam, Hemanshi Galaiya, Paul Staniland, Jessica Staniland, Ian Savill, Tuck S. Wong, and Kang L. Tee 2019. "Adaptive Laboratory Evolution of Cupriavidus necator H16 for Carbon Co-Utilization with Glycerol" International Journal of Molecular Sciences 20, no. 22: 5737. https://doi.org/10.3390/ijms20225737

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