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Development of a fluorescent nanosensor for ribose
Author(s) -
Lager Ida,
Fehr Marcus,
Frommer Wolf B,
Lalonde Sylvie
Publication year - 2003
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(03)00976-1
Subject(s) - ribose , fluorescence , periplasmic space , biochemistry , förster resonance energy transfer , chemistry , escherichia coli , monosaccharide , transporter , cell , gene , enzyme , physics , quantum mechanics
To analyze ribose uptake and metabolism in living cells, nanosensors were engineered by flanking the Escherichia coli periplasmic ribose binding protein with two green fluorescent protein variants. Following binding of ribose, fluorescence resonance energy transfer decreased with increasing ribose concentration. Five affinity mutants were generated covering binding constants between 400 nM and 11.7 mM. Analysis of nanosensor response in COS‐7 cells showed that free ribose accumulates in the cell and is slowly metabolized. Inhibitor studies suggest that uptake is mediated by a monosaccharide transporter of the GLUT family, however, ribose taken up into the cell was not or only slowly released, indicating irreversibility of uptake.