Biosynthesis of Fluorescent β Subunits of C-Phycocyanin from Spirulina subsalsa in Escherichia coli and Their Antioxidant Properties

摘要

Phycocyanin, which covalently binds phycocyanobilin chromophores, is not only a candidate fluorescent probe for biological imaging, but also a potential antioxidative agent for healthcare. Herein, a plasmid harboring two cassettes was constructed, with cpcB from Spirulina subsalsa in one cassette and the fusion gene cpcS::ho1::pcyA in the other, and then expressed in Escherichia coli. PCB-CpcB(C-82), a fluorescent phycocyanin β subunit, was biosynthesized in E. coli, exhibiting an absorption maximum at 620 nm and fluorescence emission maximum at 640 nm. When cpcS was replaced by cpcT, PCB-CpcB(C-153), another fluorescent phycocyanin β subunit, was produced, exhibiting an absorption maximum at 590 nm and fluorescence emission maximum at 620 nm. These two fluorescent biliproteins showed stronger scavenging activity toward hydroxyl and DPPH free radicals than apo-CpcB. The IC50 values for hydroxyl radical scavenging by PCB-CpcB(C-82), PCB-CpcB(C-153), and apo-CpcB were 38.72 ± 2.48 µg/mL, 51.06 ± 6.74 µg/mL, and 81.82 ± 0.67 µg/mL, respectively, and the values for DPPH radical scavenging were 201.00 ± 5.86 µg/mL, 240.34 ± 4.03 µg/mL, and 352.93 ± 26.30 µg/mL, respectively. The comparative antioxidant capacities of the proteins were PCB-CpcB(C-82) > PCB-CpcB(C-153) > apo-CpcB, due to bilin binding. The two fluorescent biliproteins exhibited a significant effect on relieving the growth of E. coli cells injured by H2O2. The results of this study suggest that the fluorescent phycocyanin β subunits of S. subsalsa were reconstructed by one expression vector in E. coli, and could be developed as potential antioxidants.