Ratiometric analyses at critical temperatures can magnify the signal intensity of FRET-based sugar sensors with periplasmic binding proteins = 분석온도 조절을 통한 당 감지 센서의 감지능 향상
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- Ratiometric analyses at critical temperatures can magnify the signal intensity of FRET-based sugar sensors with periplasmic binding proteins = 분석온도 조절을 통한 당 감지 센서의 감지능 향상
- Jongsik Gam; Jae-Seok Ha; Haseong Kim; Dae-Hee Lee; J Lee; Seung Goo Lee
- Bibliographic Citation
- Biosensors & Bioelectronics, vol. 72, no. 1, pp. 37-43
- Publication Year
- Fluorescence resonance energy transfer (FRET)-based sensors transduce ligand recognition into a change in the fluorophore spectrum, as ligand binding alters the distance between and orientation of two fluorescent proteins. Here, we report a dramatic increase in the signal intensity of FRET-based sugar sensors with bacterial periplasmic binding proteins (PBPs) in the binding moiety, by increasing the analysis temperature, usually higher than 50. °C. The increased signal intensity results from a sudden decrease in background signal at critical temperatures, while recovering the maximum FRET ratios in the presence of ligands. When tested with a maltose sensor using a maltose-binding protein as the binding moiety, the FRET ratio at the critical temperature, 55. °C, was 17-fold higher than at ambient temperatures. Similar effects were observed using analogous sensors for allose, arabinose, and glucose, providing highly dynamic and quantitative ratio changes at the critical temperatures. The proposed mechanism underlying the signal improvement is thermal relaxation of the binding proteins at the critical temperature; this hypothesis was supported by the results of intrinsic tryptophan fluorescence and circular dichroism experiments. In summary, this study shows that the conformational relaxation of proteins under specific conditions can be leveraged for highly sensitive and rapid measurements of ligands using FRET-based sensors.
- Conformation relaxationCritical temperatureMolecular biosensorFluorescence resonance energy transfer
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- Synthetic Biology and Bioengineering Research Institute > Synthetic Biology Research Center > 1. Journal Articles
Synthetic Biology and Bioengineering Research Institute > 1. Journal Articles
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