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Cited by 114biodegradable polymer synthesis and propertiesMicrobial Metabolic Engineering and BioproductionAlgal biology and biofuel production

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Photoautotrophic production of polyhydroxyalkanoates in a synthetic mixed culture of Synechococcus elongatus cscB and Pseudomonas putida cscAB

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Löwe, Hannes, Hobmeier, Karina, Moos, Manuel, Kremling, Andreas, Pflüger-Grau, Katharina·2017·Biotechnology for Biofuels
DOI 10.1186/s13068-017-0875-0PMC PMC5517840
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Abstract

One of the major challenges for the present and future generations is to find suitable substitutes for the fossil resources we rely on today. Cyanobacterial carbohydrates have been discussed as an emerging renewable feedstock in industrial biotechnology for the production of fuels and chemicals, showing promising production rates when compared to crop-based feedstock. However, intrinsic capacities of cyanobacteria to produce biotechnological compounds are limited and yields are low. Here, we present an approach to circumvent these problems by employing a synthetic bacterial co-culture for the carbon-neutral production of polyhydroxyalkanoates (PHAs) from CO 2 . The co-culture consists of two bio - modules : Bio - module I , in which the cyanobacterial strain Synechococcus elongatus cscB fixes CO 2 , converts it to sucrose, and exports it into the culture supernatant; and bio - module II , where this sugar serves as C-source for Pseudomonas putida cscAB and is converted to PHAs that are accumulated in the cytoplasm. By applying a nitrogen-limited process, we achieved a maximal PHA production rate of 23.8 mg/(L day) and a maximal titer of 156 mg/L. We will discuss the present shortcomings of the process and show the potential for future improvement. These results demonstrate the feasibility of mixed cultures of S. elongatus cscB and P. putida cscAB for PHA production, making room for the cornucopia of possible products that are described for P. putida . The construction of more efficient sucrose-utilizing P. putida phenotypes and the optimization of process conditions will increase yields and productivities and eventually close the gap in the contemporary process. In the long term, the co-culture may serve as a platform process, in which P. putida is used as a chassis for the implementation of synthetic metabolic pathways for biotechnological production of value-added products.

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Crossref · 10.1186/s13068-017-0875-0

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Fig. 1 Concept of the synthetic co-culture of S. elongatus cscB and P. putida cscAB for the production of PHA from CO2 and light. CO2 is fixed via the Calvin cycle to make sucrose, which in turn is secreted into the surrounding medium by the activity of the heterologous sucrose permease CscB. CscA produced by P. putida cscAB leaks out of the cell, where it splits sucrose extracellularly [16]. The monomers (glucose and fructose) are metabolized by P. putida cscAB, and polyhydroxyalkanoates (PHA) are accumulated in the cytoplasm [15]
Fig. 1 Concept of the synthetic co-culture of S. elongatus cscB and P. putida cscAB for the production of PHA from CO2 and light. CO2 is fixed via the Calvin cycle to make sucrose, which in turn is secreted into the surrounding medium by the activity of the heterologous sucrose permease CscB. CscA produced by P. putida cscAB leaks out of the cell, where it splits sucrose extracellularly [16]. The monomers (glucose and fructose) are metabolized by P. putida cscAB, and polyhydroxyalkanoates (PHA) are accumulated in the cytoplasm [15]CC BY · PubMed Central
Fig. 2 Growth and sucrose concentration from fermentations of S. elongatus cscB in a photobioreactor in BG-11+ in the presence of different salt concentrations. Cells were grown in BG11+ medium with the NaCl concentration indicated, and growth and sucrose secretion were monitored. CscB production was induced with 0.1 mM IPTG at an OD of 0.1–0.2
Fig. 2 Growth and sucrose concentration from fermentations of S. elongatus cscB in a photobioreactor in BG-11+ in the presence of different salt concentrations. Cells were grown in BG11+ medium with the NaCl concentration indicated, and growth and sucrose secretion were monitored. CscB production was induced with 0.1 mM IPTG at an OD of 0.1–0.2CC BY · PubMed Central
Fig. 3 Growth of P. putida cscAB in modified cyanobacterial BG-11 medium supplemented with glucose, fructose (each 1.5 g/L), and 150 mM NaCl. Experiments were conducted in 250 mL unbaffled shake flasks with an effective volume of 25 mL. Temperature was set to 30 °C, and agitation rate to 220 rpm. Shown are the means and standard deviations calculated from three biological replicates
Fig. 3 Growth of P. putida cscAB in modified cyanobacterial BG-11 medium supplemented with glucose, fructose (each 1.5 g/L), and 150 mM NaCl. Experiments were conducted in 250 mL unbaffled shake flasks with an effective volume of 25 mL. Temperature was set to 30 °C, and agitation rate to 220 rpm. Shown are the means and standard deviations calculated from three biological replicatesCC BY · PubMed Central
Fig. 4 Co-cultivation of S. elongatus cscB and P. putida cscAB in the photobioreactor. Optical density, Nile red-stained cell count of P. putida cscAB, cell count of S. elongates cscB and colony forming units (CFU) determined from plating are depicted over time in a. Arrows indicate the induction of sucrose export by the addition of 0.1 mM IPTG and inoculation of P. putida cscAB. Sucrose and PHA concentrations are plotted in b. Uncertainties in PHA concentrations were estimated from the propagation of errors of the PHA standards
Fig. 4 Co-cultivation of S. elongatus cscB and P. putida cscAB in the photobioreactor. Optical density, Nile red-stained cell count of P. putida cscAB, cell count of S. elongates cscB and colony forming units (CFU) determined from plating are depicted over time in a. Arrows indicate the induction of sucrose export by the addition of 0.1 mM IPTG and inoculation of P. putida cscAB. Sucrose and PHA concentrations are plotted in b. Uncertainties in PHA concentrations were estimated from the propagation of errors of the PHA standardsCC BY · PubMed Central
Fig. 5 Nitrate-limited co-cultivation of S. elongatus cscB and P. putida cscAB. Optical density, Nile red-stained cell count of P. putida cscAB and cell count of S. elongatus cscB are depicted over time in a. Sucrose, PHA and nitrate concentrations are plotted in b. Uncertainties in PHA concentrations were estimated from the propagation of errors of the PHA standards. Nitrate was fed at a constant rate of 46 ± 2 mg/day after inoculation with P. putida cscAB
Fig. 5 Nitrate-limited co-cultivation of S. elongatus cscB and P. putida cscAB. Optical density, Nile red-stained cell count of P. putida cscAB and cell count of S. elongatus cscB are depicted over time in a. Sucrose, PHA and nitrate concentrations are plotted in b. Uncertainties in PHA concentrations were estimated from the propagation of errors of the PHA standards. Nitrate was fed at a constant rate of 46 ± 2 mg/day after inoculation with P. putida cscABCC BY · PubMed Central

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Metadata

Authors
Löwe, H., Hobmeier, K., Moos, M., Kremling, A., & Pflüger-Grau, K.
Affiliations
Technical University of Munich (DE)
ORCID
Venue
Biotechnology for Biofuels
Publisher
Springer Science and Business Media LLC
Published
2017-07-19
Volume
10
Issue
1
Pages
190-190
ISSN
1754-6834 (ISSN-L)
DOI
10.1186/s13068-017-0875-0
PMC
PMC5517840
OpenAlex
W2736712864
Cited by
114
References
35
Citations by year
2026: 6 · 2025: 10 · 2024: 13 · 2023: 15 · 2022: 20 · 2021: 18 · 2020: 21 · 2019: 3 · 2018: 7 · 2017: 1
FWCI
3.15
Type
journal article
Language
en
Open access
Licence
cc-by
Locations
  • Biotechnology for Biofuels (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • mediaTUM – the media and publications repository of the Technical University Munich (Technical University Munich) (submittedVersion, other-oa)
Funders
  • U.S. Department of Defense
  • Technische Universität München
  • Centro Nacional de Biotecnología
Topics
  • Physical Sciences › Materials Science › Biomaterials › biodegradable polymer synthesis and properties
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Microbial Metabolic Engineering and Bioproduction
  • Physical Sciences › Energy › Renewable Energy, Sustainability and the Environment › Algal biology and biofuel production
Keywords
polyhydroxyalkanoates, sucrose export, renewable raw materials, biopolymer production, chassis strain, synthetic metabolic pathways

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