Assessing the potential of sucrose production by genetically engineered cyanobacteria
Zusammenfassung der Projektergebnisse
During the last years, cyanobacteria were applied for the production of different biofuels, however, in most cases the productivity was rather low. Our project aimed to generate sucrose-producing strains of cyanobacteria for the future CO2-neutral production of biochemicals and feedstock. Cyanobacteria are naturally competent to synthesize sucrose, since this sugar is accumulated by many strains as compatible solute under salt stress. Sucrose is an attractive product, because it can be directly used for nutrition or as carbon source for heterotrophic bacteria, which are producing high value products in biotechnology. To support such future application, the production of sucrose needs to be enhanced, which can be achieved by the knock-out of competing pathways or by introducing a strong sucrose sink due to the continuous export of the product into the medium. The project was carried out in cooperation with the team of Prof. Xuefeng Lu, Qingdao Institute of Bioenergy and Bioprocess Technology, Microbial Metabolic Engineering Group, Chinese Academy of Sciences, Qingdao, China. During the project we concentrated our work on the model strain Synechocystis sp. PCC 6803. It produces sucrose as secondary compatible solute. From Synechocystis exist several substrains; the work was carried out with the glucose-tolerant strain that showed the highest natural sucrose accumulation. First, we identified enzymes, which potentially cleave the compatible solutes sucrose and glucosylglycerol (GG). The invertase-encoding gene was identified in Synechocystis and its function was verified by biochemical characterization of the recombinant protein. Knock-out of invertase at least doubled the salt-induced sucrose accumulation in all sucrose-producing strain backgrounds. Interestingly, the invertase mutant still represents the only strain, which accumulates sucrose already under NaCl-free conditions. For GG, we also identified and verified a GG-hydrolase, which plays an important role during the acclimation of Synechocystis to hypo-osmotic shock, but its inactivation had not influence on GG steady state levels. Second, we inactivated GG synthesis as competing pool for sucrose accumulation, which resulted in another twofold increased sucrose-pool, whereas the combined knock-out of invertase and GG synthesis resulted in 10fold more sucrose compared to wild type. Third, glycogen synthesis was abolished as competing pool, but these strains showed diminished growth and stress tolerance, which clearly indicated that rather glycogen breakdown but not its synthesis should be targeted in future attempts to enhance sucrose accumulation. Forth, we identified plant sucrose exporters such as SWEET11, which support efficient sucrose export from Synechocystis, which resulted in the highest sucrose productivity. Summarizing, a great number of improved sucrose-producing Synechocystis strains was generated. These strains could be used for the future biotechnological application. Moreover, these strains also provide an excellent tool box to study the impact of organic carbon sinks such as sucrose on cyanobacterial photosynthesis and carbon allocation.
Projektbezogene Publikationen (Auswahl)
- 2015. Salt acclimation of cyanobacteria and their application in biotechnology. Life 5:25-49
Nadin Pade, Martin Hagemann
(Siehe online unter https://dx.doi.org/10.3390/life5010025) - 2017. The glucosylglycerol degrading enzyme GghA is involved in the acclimation to fluctuating salinities of the cyanobacterium Synechocystis sp. strain PCC 6803. Microbiology 163:1319-1328
Friedrich Kirsch, Nadin Pade, Stephan Klähn, Wolfgang R. Hess, Martin Hagemann
(Siehe online unter https://doi.org/10.1099/mic.0.000518) - 2017. The response regulator Slr1588 regulates spsA but is not crucial for salt acclimation of Synechocystis sp. PCC 6803. Frontiers in Microbiology 8:1176
Kuo Song, Martin Hagemann, Xiaoming Tan, Xuefeng Lu
(Siehe online unter https://doi.org/10.3389/fmicb.2017.01176) - 2018. Effects of reduced and enhanced glycogen pools on salt-induced sucrose production in a sucrose-secreting strain of Synechococcus elongatus PCC 7942. Applied and Environmental Microbiology 84:e02023-17
Cuncun Qiao, Yangkai Duan, Mingyi Zhang, Martin Hagemann, Quan Luo, Xuefeng Lu
(Siehe online unter https://doi.org/10.1128/AEM.02023-17) - 2018. Systems and synthetic biology for the biotechnological application of cyanobacteria. Current Opinion in Biotechnology 49:94-99
Martin Hagemann, Wolfgang R. Hess
(Siehe online unter https://doi.org/10.1016/j.copbio.2017.07.008)
