FATTY ACID AND LIPID LEVELS IN CYANOBACTERIUM Synechocystis sp. PCC 6803 MUTANT OVEREXPRESSING plsX GENE

Abstract

In this study, we successfully constructed the plsX- overexpressing strain of Synechocystis sp. PCC 6803 (OX_plsX strain), and further studied the effects of salt stress and nutrient modifications on growth, pigment contents of chlorophyll a and carotenoids, oxygen evolution rate or photosynthetic efficiency, lipid content and fatty acid composition, and transcript level of genes related to fatty acid synthesis compared to Synechocystis wild type. Initially, both strains were cultured in normal BG11 medium. Cell growth and intracellular pigment contents of OX_plsX strain were not different from those of wild type, as well as their oxygen evolution rate. However, both total lipid and unsaturated lipid contents of OX_plsX strain were higher than those of wild type. After that, effect of salt stress and nutrient modifications on cell lipid and fatty acid production were studied. It was found that sodium chloride at 100 mM concentration could induce the accumulation of total lipid in OX_plsX strain higher than wild type, up to about 1.3 fold whereas unsaturated lipid content of OX_plsX strain was higher than wild type in all salt stress studied. The fatty acid composition from GC-MS analysis consisted of saturated and unsaturated fatty acids including palmitic acid (C16:0), stearic acid (C18:0), pailmitoleic acid (C16:1), oleic acid (C18:1), linoleic acid (C18:2) and α-linolenic acid (C18:3). We found that salt stress effectively influenced the intracellular composition ratio of saturated and unsaturated fatty acids. On the other hand, after transferring log-phase growing cells into adapted BG11 medium, with nitrogen deficiency or carbon source supplementation of 0.4% acetate (6.7 mM concentration), acetate supplementation enabled OX_plsX strain to accumulate more total lipids and unsaturated lipids of about 1.3 and 2.9 fold, respectively, when compared to wild type. Next, we aligned the amino sequence of Synechocystis plsX compared with the amino sequence of cyanobacterium Anabaena sp. PCC 7120 and Synechococcus sp. PCC 7002 PlsXs. They showed high identity at 65.77% and 72.70%, respectively, whereas the identity comparing with plant Arabidopsis thaliana glycerol-3-phosphate acyltransferase was about 16.09%. The identity between Synechocystis PlsX and bacterial Escherichia coli PlsX and Bacillus subtilis PlsX were 36.72% and 42.42%, respectively. Phylogenetic tree analysis suggests that Synechocystis plsX may be classified into phosphate acyltransferase enzyme Altogether, plsX gene had a crucial role in fatty acid synthesis since the overexpression of plsX gene enhanced the lipid accumulation in Synechocystis cells. Also, the acetate supplementation could induce the lipid production in both wild type and plsX-overexpressing strains.