The Effects of Salts and Osmoprotectants on Enzyme Activities of Fructose-1,6-biphosphate Aldolases in a Halotolerant Cyanobacterium, Halothece sp. PCC 7418

The halotolerant cyanobacterium, Halothece sp. PCC 7418, possesses two classes of fructose-1,6-bisphosphate aldolase (FBA): H2846 and H2847. Though class I (CI)-FBA H2846 is thought to be associated with salt tolerance, the regulatory mechanisms, molecular characteristics, and expression profiles between H2846 and class II (CII)-FBA H2847 have scarcely been investigated. Here, we show that the accumulation of the H2846 protein is highly responsive to both up- and down-shock with NaCl, whereas H2847 is constitutively expressed. The activity of CI- and CII-FBA in cyanobacterial extracts is correlated with the accumulation patterns of H2846 and H2847, respectively. In addition, it was found that these activities were inhibited by NaCl and KCl, with CII-FBA activity strikingly inhibited. It was also found that the CI-FBA activity of recombinant H2846 was hindered by salts and that this hindrance could be moderated by the addition of glycine betaine (GB), whereas no moderation occurred with other potential osmoprotectant molecules (proline, sucrose, and glycerol). In addition, a phylogenetic analysis showed that CI-FBAs with higher similarities to H2846 tended to be distributed among potential GB-synthesizing cyanobacteria. Taken together, our results provide insights into the independent evolution of the CI- and CII-FBA gene families, which show distinct expression profiles and functions following salt stress.

Induction of Antioxidative Activity and Antioxidant Molecules in the Halotolerant Cyanobacterium Halothece sp. PCC7418 by Temperature Shift

Extremophiles are a rich source of novel secondary metabolites and valuable antioxidative compounds. Here, we examined the antioxidative capacities of aqueous extracts derived from the halotolerant cyanobacterium Halothece sp. PCC7418. The antioxidative activities of extracts derived from Halothece sp. PCC7418 cells exposed to temperature shock were significantly enhanced. Quantification of antioxidant molecules (phenolic compounds and phycobiliproteins) in the aqueous extracts revealed that the amounts of these molecules were modulated by temperature shock. In addition, the intracellular accumulation of mycosporine-2-glycine, a potential antioxidative molecule, was strongly enhanced by cold shock. Our results show that the treatment of Halothece sp. PCC7418 cells with temperature shock may allow for the robust production of antioxidants.

Efficient Bioproduction of Mycosporine-2-glycine, Which Functions as Potential Osmoprotectant, using Escherichia coli Cells

Mycosporine-2-glycine (M2G) is known to be synthesized in halotolerant cyanobacterium Aphanothece halophytica. Escherichia coli cells in which the M2G synthetic genes of A. halophytica were introduced could synthesize M2G. Here, we report that M2G producing transformed E. coli cells showed salt tolerance compared to control cells. This result suggested that M2G could function as a potential osmoprotectant in E. coli. To our knowledge, this is the first report presenting the evidence that mycosporine-like amino acid confers salt tolerance on E. coli. Intracellular M2G content in the transformed E. coli cells were varied depending on NaCl concentration with maximum level at 0.75 M. Moreover, intracellular M2G level was affected by a supply of glycine with maximum level at 5 mM. In conclusion, we found that transformed E. coli cells could produce 205 μg of M2G/g fresh weight of cells under the best effective growth condition in this study. Thus, the results obtained here offer the potential for the bioproduction of mycosporine-like amino acids using the genetically engineered E. coli cells.