Carbon flux to growth or polyhydroxyalkanoate synthesis under microaerophilic conditions is affected by fatty acid chain-length in Pseudomonas putida LS46
- 204 Downloads
Economical production of medium-chain length polyhydroxyalkanoates (mcl-PHA) is dependent on efficient cultivation processes. This work describes growth and mcl-PHA synthesis characteristics of Pseudomonas putida LS46 when grown on medium-chain length fatty acids (octanoic acid) and lower-cost long-chain fatty acids (LCFAs, derived from hydrolyzed canola oil) in microaerophilic environments. Growth on octanoic acid ceased when the oxygen uptake rate was limited by the oxygen transfer rate, and mcl-PHA accumulated to 61.9% of the cell dry mass. From LCFAs, production of non-PHA cell mass continued at a rate of 0.36 g L−1 h−1 under oxygen-limited conditions, while mcl-PHA accumulated simultaneously to 31% of the cell dry mass. The titer of non-PHA cell mass from LCFAs at 14 h post-inoculation was double that obtained from octanoic acid in bioreactors operated with identical feeding and aeration conditions. While the productivity for octanoic acid was higher by 14 h, prolonged cultivation on LCFAs achieved similar productivity but with twice the PHA titer. Simultaneous co-feeding of each substrate demonstrated the continued cell growth under microaerophilic conditions characteristic of LCFAs, and the resulting polymer was dominant in C8 monomers. Furthermore, co-feeding resulted in improved PHA titer and volumetric productivity compared to either substrate individually. These results suggest that LCFAs improve growth of P. putida in oxygen-limited environments and could reduce production costs since more non-PHA cell mass, the cellular factories required to produce mcl-PHA and the most oxygen-intensive cellular process, can be produced for a given oxygen transfer rate.
KeywordsPHA Dissolved oxygen Bioreactor Biopolymer Fed-batch LCFAs
This study was funded by the Genome Canada, through the Genome Applications and Partnership Program (GAPP), and the Natural Sciences and Engineering Research Council (NSERC) of Canada through a Collaborative Research and Development (CRD) grant with Minto BioProducts Ltd. as the industrial partner (grant number CRDPJ-490630-15).
Compliance with ethical standards
Conflict of interest
The authors declare they have no conflict of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
- Andin N, Longieras A, Veronese T, Marcato F, Molina-Jouve C, Uribelarrea JL (2017) Improving carbon and energy distribution by coupling growth and medium chain length polyhydroxyalkanoate production from fatty acids by Pseudomonas putida KT2440. Biotechnol Bioprocess Eng 22:308–318. https://doi.org/10.1007/s12257-016-0449-1 CrossRefGoogle Scholar
- Blunt W, Dartiailh C, Sparling R, Gapes D, Levin DB, Cicek N (2017) Microaerophilic environments improve the productivity of medium chain length polyhydroxyalkanoate biosynthesis from fatty acids in Pseudomonas putida LS46. Process Biochem 59:18–25. https://doi.org/10.1016/j.procbio.2017.04.028 CrossRefGoogle Scholar
- Davis R, Duane G, Kenny ST, Cerrone F, Guzik MW, Babu RP, Casey E, O’Connor KE (2015) High cell density cultivation of Pseudomonas putida KT2440 using glucose without the need for oxygen enriched air supply. Biotechnol Bioeng 112:725–733. https://doi.org/10.1002/bit.25474 CrossRefPubMedGoogle Scholar
- Fernández D, Rodríguez E, Bassas M, Viñas M, Solanas AM, Llorens J, Marqués AM, Manresa A (2005) Agro-industrial oily wastes as substrates for PHA production by the new strain Pseudomonas aeruginosa NCIB 40045: effect of culture conditions. Biochem Eng J 26:159–167. https://doi.org/10.1016/j.bej.2005.04.022 CrossRefGoogle Scholar
- Fontaine P, Mosrati R, Corroler D (2017) Medium chain length polyhydroxyalkanoates biosynthesis in Pseudomonas putida mt-2 is enhanced by co-metabolism of glycerol/octanoate or fatty acids mixtures. Int J Biol Macromol 98:430–435. https://doi.org/10.1016/j.ijbiomac.2017.01.115 CrossRefPubMedGoogle Scholar
- Grousseau E, Blanchet E, Déléris S, Albuquerque MGE, Paul E, Uribelarrea JL (2013) Impact of sustaining a controlled residual growth on polyhydroxybutyrate yield and production kinetics in Cupriavidus necator. Bioresour Technol 148:30–38. https://doi.org/10.1016/j.biortech.2013.08.120 CrossRefPubMedGoogle Scholar
- Haba E, Vidal-Mas J, Bassas M, Espuny MJ, Llorens J, Manresa A (2007) Poly 3-(hydroxyalkanoates) produced from oily substrates by Pseudomonas aeruginosa 47T2 (NCBIM 40044): effect of nutrients and incubation temperature on polymer composition. Biochem Eng J 35:99–106. https://doi.org/10.1016/j.bej.2006.11.021 CrossRefGoogle Scholar
- Kellerhals MB, Kessler B, Witholt B, Tchouboukov A, Brandl H (2000) Renewable long-chain fatty acids for production of biodegradable medium-chain-length polyhydroxyalkanoates (mcl-PHAs) at laboratory and pilot plant scales. Macromolecules 33:4690–4698. https://doi.org/10.1021/ma000655k CrossRefGoogle Scholar
- Kim GJ, Lee IY, Yoon SC, Shin YC, Park YH (1997) Enhanced yield and a high production of medium-chain-length poly(3-hydroxyalkanoates) in a two-step fed-batch cultivation of Pseudomonas putida by combined use of glucose and octanoate. Enzym Microb Technol 20:500–505. https://doi.org/10.1016/S0141-0229(96)00179-2 CrossRefGoogle Scholar
- Kurth N, Brachet F, Robic D, Bourbouze R, Renard E, Guerin P (2001) Poly(3-hydroxyoctanoate) containing pendant carboxylic groups for the preparation of nanoparticles aimed at drug transport and release. Polymer (Guildf) 43:1095–1101. https://doi.org/10.1016/S0032-3861(01)00692-9 CrossRefGoogle Scholar
- Lee SY, Wong HH, Il CJ, Lee SH, Lee SC, Han CS (2000) Production of medium-chain-length polyhydroxyalkanoates by high-cell- density cultivation Pseudomonas putida under phosphorus limitation. Biotechnol Bioeng 68:466–470. https://doi.org/10.1002/(SICI)1097-0290(20000520)68:4<466::AID-BIT12>3.0.CO;2-T CrossRefPubMedGoogle Scholar
- Nikodinovic-Runic J, Guzik M, Kenny ST, Babu R, Werker A, O’Connor KE (2013) Carbon-rich wastes as feedstocks for biodegradable polymer (polyhydroxyalkanoate) production using bacteria. Adv Appl Microbiol 84:139–200. https://doi.org/10.1016/B978-0-12-407673-0.00004-7 CrossRefPubMedGoogle Scholar