Rerouting of carbon flux in a glycogen mutant of cyanobacteria assessed via isotopically non-stationary 13C metabolic flux analysis

4 years ago

Rerouting of carbon flux in a glycogen mutant of cyanobacteria assessed via isotopically non-stationary 13C metabolic flux analysis

John I. Hendry, Niels-Ulrik Frigaard, Doug K. Allen, K. Benedikt Möllers, Madhuri Digmurti, Fangfang Ma, Damini Jaiswal, Charulata Prasannan, Santanu Dasgupta, Pramod P. Wangikar

Cyanobacteria, which constitute a quantitatively dominant phylum, have attracted attention in biofuel applications due to favorable physiological characteristics, high photosynthetic efficiency and amenability to genetic manipulations. However, quantitative aspects of cyanobacterial metabolism have received limited attention. In the present study, we have performed isotopically non-stationary 13C metabolic flux analysis (INST-13C-MFA) to analyze rerouting of carbon in a glycogen synthase deficient mutant strain (glgA-I glgA-II) of the model cyanobacterium Synechococcus sp. PCC 7002. During balanced photoautotrophic growth, 10–20% of the fixed carbon is stored in the form of glycogen via a pathway that is conserved across the cyanobacterial phylum. Our results show that deletion of glycogen synthase gene orchestrates cascading effects on carbon distribution in various parts of the metabolic network. Carbon that was originally destined to be incorporated into glycogen gets partially diverted toward alternate storage molecules such as glucosylglycerol and sucrose. The rest is partitioned within the metabolic network, primarily via glycolysis and tricarboxylic acid cycle. A lowered flux toward carbohydrate synthesis and an altered distribution at the glucose-1-phosphate node indicate flexibility in the network. Further, reversibility of glycogen biosynthesis reactions points toward the presence of futile cycles. Similar redistribution of carbon was also predicted by Flux Balance Analysis. The results are significant to metabolic engineering efforts with cyanobacteria where fixed carbon needs to be re-routed to products of interest. Biotechnol. Bioeng. 2017;9999: 1–11. © 2017 Wiley Periodicals, Inc. INST-13C-MFA study of a Synechococcus sp PCC 7002 mutant lacking the enzyme glycogen synthase showed that sucrose and glucosyl-glycerol are the major alternative carbon sinks in this mutant. Significant reduction in the flux through phosphoglucose isomerase and phosphoglucomutase reactions in the mutant indicates an overall reduction in the flow of carbon towards the synthesis of glucose-1-phosphate. The shift in the UDP-glucose pyrophosphorylase to ADP-glucose pyrophosphorylase flux ratio in the mutant demonstrates the flexibility of the glucose-1-phosphate branch point.

Publisher URL: http://onlinelibrary.wiley.com/resolve/doi

DOI: 10.1002/bit.26350