SE179:/MS01

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Sample Set Information

ID	TSE1337
Title	Genetic manipulation of a metabolic enzyme and a transcriptional regulator increasing succinate excretion from unicellular cyanobacterium.
Description	Succinate is a building block compound that the U.S. Department of Energy (DOE) has declared as important in biorefineries, and it is widely used as a commodity chemical. Here, we identified the two genes increasing succinate production of the unicellular cyanobacterium Synechocystis sp. PCC 6803. Succinate was excreted under dark, anaerobic conditions, and its production level increased by knocking out ackA, which encodes an acetate kinase, and by overexpressing sigE, which encodes an RNA polymerase sigma factor. Glycogen catabolism and organic acid biosynthesis were enhanced in the mutant lacking ackA and overexpressing sigE, leading to an increase in succinate production reaching five times of the wild-type levels. Our genetic and metabolomic analyses thus demonstrated the effect of genetic manipulation of a metabolic enzyme and a transcriptional regulator on succinate excretion from this cyanobacterium with the data based on metabolomic technique.
Authors	Osanai, T., Shirai, T., Iijima, H., Nakaya, Y., Okamoto, M., Kondo, A. and Hirai, M.Y.
Reference	Front Microbiol. 2015 Oct 6;6:1064. doi: 10.3389/fmicb.2015.01064.
Comment

ID	MS01
Title	LC-MS/MS analysis
Instrument	LCMS‐8040 triple quadrupole LC/MS/MS spectrometer; Shimadzu
Instrument Type
Ionization	ESI
Ion Mode
Description	Equal amounts of cells (10 mL cell culture with A730 = 1.0) were harvested by rapid filtration, and metabolites were extracted using a previously described method (Osanai et al., 2014b). Briefly, the cells were filtrated, and then the intermediate metabolites were quenched and extracted in 1.2 mL of solvent mixture (CHCl3:CH3OH:H2O, 2.5:2.5:1, v/v/v) containing 10 μg/L D-(+)-camphor-10-sulfonic acid as an internal standard. After centrifugation at 15,000 × g at 4°C for 5 min, 400 μL of the upper phase was transferred to a new tube and vacuum-dried.
Comment_of_details