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Enhanced growth at low light intensity in the cyanobacterium Synechocystis PCC 6803 by overexpressing phosphoenolpyruvate carboxylase
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Molecular Biomimetics.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Molecular Biomimetics. Hasanuddin Univ, Fac Marine Sci & Fisheries, Jl Perintis Kemerdekaan Km 10, Makassar, South Sulawesi, Indonesia..
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - Ångström, Molecular Biomimetics.
2016 (English)In: ALGAL RESEARCH-BIOMASS BIOFUELS AND BIOPRODUCTS, ISSN 2211-9264, Vol. 16, p. 275-281Article in journal (Refereed) Published
Resource type
Text
Abstract [en]

Synechocystis PCC 6803 strains overexpressing pepc, gene encoding the carbon fixing enzyme phosphoenolpyruvate carboxylase (PEPc), were constructed and characterized for growth, PEPc protein content and in vitro PEPc activities. Synechocystis strains WT + Km(r) - one (native) copy of pepc (control), WT + 2xPEPc - native copy of pepc and two additional native copies of pepc (in total three copies of pepc), and WT + PPM - native copies of ppsa (encoding phosphoenolpyruvate synthase), pepc and mdh (encoding malate dehydrogenase) and one additional copy of each gene (in total two copies each of ppsa, pepc and mdh) were analyzed for growth under normal and low light intensities, and in darkness (no growth). No significant differences in the growth rates were observed when the cells were grown under normal light intensity. However, growth under low light intensity (3 mu mol photons.m(-2).sec(-1)) resulted in increased growth rate, in particular in the strain with 3 copies of pepc. SDS-PAGE/Western immunoblots using antibodies directed against PEPc demonstrated an increased level of PEPc protein with increasing number of copies of pepc. This was followed by increased levels of in vitro PEPc activities. A less efficient ribulose 1,5-bisphosphate carboxylase/oxygenase in combination with reduced levels of NADPH and ATP under low light condition may make the relatively more efficient carbon fixing enzyme PEPc the limiting step for growth under this condition.

Place, publisher, year, edition, pages
2016. Vol. 16, p. 275-281
Keywords [en]
Genetic engineering, Enhanced growth, pepc, Phosphoenolpyruvate carboxylase, Synechocystis
National Category
Bio Materials
Identifiers
URN: urn:nbn:se:uu:diva-297764DOI: 10.1016/j.algal.2016.03.027ISI: 000375610000032OAI: oai:DiVA.org:uu-297764DiVA, id: diva2:943679
Funder
Knut and Alice Wallenberg Foundation, 2011.0067Swedish Energy AgencyAvailable from: 2016-06-28 Created: 2016-06-28 Last updated: 2019-09-01Bibliographically approved
In thesis
1. Increased Carbon Fixation for Chemical Production in Cyanobacteria
Open this publication in new window or tab >>Increased Carbon Fixation for Chemical Production in Cyanobacteria
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The combustion of fossil fuels has created many environmental problems, the major one, the greenhouse effect. Thus, we need solutions in order to replace fossil fuels and recycle the CO2 in the atmosphere. Renewable energies have created attention the last decades but electricity is the main energy form obtained. Photosynthetic organisms (including cyanobacteria) can be used as cell factories since they can convert solar energy to chemical energy. In addition, the requisites to grow them are few; light water, CO2 and inorganic nutrients. Cyanobacteria have been genetically engineered in order to produce numerous chemicals and fuels of human interest in direct processes. However, the amount of product obtained is still low. Increased carbon fixation in cyanobacteria results in higher production of carbon-based substances. This thesis focuses on the effects of overexpressing the native phosphoenolpyruvate carboxylase (PEPc) in the model cyanobacterium Synechocystis PCC 6803. PEPc is an essential enzyme and provides oxaloacetate, an intermediate of the tricarboxylic acid cycle (TCA cycle). The TCA cycle is involved in connecting the carbon and nitrogen metabolism in cyanobacteria. The strains were further engineered to produce ethylene and succinate, two examples of interests for the chemical and fuel industry. Strains with additional PEPc produced significantly more ethylene and succinate. Moreover, an in vitro characterization of PEPc from the cyanobacterium Synechococcus PCC 7002 was performed. The focus was on oligomerization state, kinetics and the structure of the carboxylase. This thesis demonstrates that increasing carbon fixation and discovering the bottlenecks in chemical production can lead to higher yields and gives us hope that cyanobacteria can be commercialized.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2019. p. 65
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1848
National Category
Engineering and Technology
Identifiers
urn:nbn:se:uu:diva-392234 (URN)978-91-513-0736-7 (ISBN)
Public defence
2019-10-18, Häggsalen, Ångströmlaboratoriet, Lägerhyddsvägen 1, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2019-09-25 Created: 2019-09-01 Last updated: 2019-10-15

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Durall de la Fuente, ClaudiaLindblad, Peter

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