A fluorescence resonance energy transfer sensor based on maltose binding protein.

A fluorescence resonance energy-transfer (FRET) sensing system for maltose based on E. coli maltose binding protein (MBP) is demonstrated. The FRET donor portion of the sensing system consists of MBP modified with long wavelength-excitable cyanine dyes (Cy3 or Cy3.5). The novel acceptor portion of the sensor consists of β-cyclodextrin (β-CD) modified with either the cyanine dye Cy5 or the dark quencher QSY9. Binding of the modified β-CD to dye-conjugated MBP results in assembly of the FRET complex. Added maltose displaces the β-CD−dye adduct and disrupts the FRET complex, resulting in a direct change in fluorescence of the donor moiety. In the use of these FRET pairs, MBP dissociation values for maltose were estimated (0.14−2.90 μM). Maltose limits of detection were in the 50−100 nm range.