Abstract
BackgroundRecent advances in the understanding of photosensing in biological systems have enabled the use of photoreceptors as novel genetic tools. Exploiting various photoreceptors that cyanobacteria possess, a green light-inducible gene expression system was previously developed for the regulation of gene expression in cyanobacteria. However, the applications of cyanobacterial photoreceptors are not limited to these bacteria but are also available for non-photosynthetic microorganisms by the coexpression of a cyanobacterial chromophore with a cyanobacteria-derived photosensing system. An Escherichia coli-derived self-aggregation system based on Antigen 43 (Ag43) has been shown to induce cell self-aggregation of various bacteria by exogenous introduction of the Ag43 gene.ResultsAn E. coli transformant harboring a plasmid encoding the Ag43 structural gene under a green light-regulated gene expression system derived from the cyanobacterium Synechocystis sp. PCC6803 was constructed. Ag43 was inserted downstream of the cpcG2 promoter PcpcG2, and its expression was regulated by green light induction, which was achieved by the functional expression of cyanobacterial CcaS/CcaR by coexpressing its chromophore synthesis gene cassette in E. coli. E. coli transformants harboring this designed system self-aggregated under green light exposure and precipitated, whereas transformants lacking the green light induction system did not. The green light induction system effectively functioned before the cell culture entered the stationary growth phase, and approximately 80 % of the cell culture was recovered by simple decantation.ConclusionThis study demonstrated the construction of a cell recovery system for non-photosynthetic microorganisms induced by exposure of cells to green light. The system was regulated by a two-component regulatory system from cyanobacteria, and cell precipitation was mediated by an autotransporter protein, Ag43. Although further strict control and an increase of cell recovery efficiency are necessary, the system represents a novel tool for future bioprocessing with reduced energy and labor required for cell recovery.
Highlights
Recent advances in the understanding of photosensing in biological systems have enabled the use of photoreceptors as novel genetic tools
The Antigen 43 (Ag43) structural gene was inserted downstream of the cpcG2 promoter, PcpcG2, and its expression was regulated by green light induction, achieved by the functional expression of cyanobacterial CcaS/CcaR by coexpression of its chromophore synthesis gene cassette in E. coli
Transformants harboring pSTVPCB and pBRGLAg∆S, pBRGLAg∆R, or pBRGLAg∆SR, cultured under red or green light, showed no decrease in OD595 or OD600, suggesting that these transformants had no precipitation ability (Fig. 2b, c, d). These results indicated that cell precipitation ability was induced when the transformants harbored both genes, those for PCB synthesis and for the complete two-component regulatory system CcaS/CcaR, and only when cultured under green light
Summary
Recent advances in the understanding of photosensing in biological systems have enabled the use of photoreceptors as novel genetic tools. Exploiting various photoreceptors that cyanobacteria possess, a green lightinducible gene expression system was previously developed for the regulation of gene expression in cyanobacteria. Recent advances in the understanding of photosensing in biological systems have permitted the use of photoreceptors as novel genetic tools [1,2,3,4,5,6,7,8]. Cyanobacteria have various light-sensing systems to effectively regulate photosynthesis [9, 10] and avoid photo-inhibition caused by strong or short-wavelength light [11,12,13,14,15,16,17]. By exploiting various photoreceptors in cyanobacteria, a green light-inducible gene expression system has been developed.
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