Principles of Fluorescence Spectroscopy

PRINCIPLES OF FLUORESCENCE SPECTROSCOPY 653
Figure 19.59. Schematic of a surface-bound maltose sensor based on the maltose-binding protein from E. coli and RET. Reprinted with permission
from [220]. Copyright © 2004, American Chemical Society.
because the changes in distance have not been large enough
to result in large changes in the transfer efficiency.
This problem of a limited change in intensity was
solved by the use of RET between surface-bound reagents
(Figure 19.59). This example also illustrates the increasingly
sophisticated chemistry of sensors. This maltose sensor is
based on the maltose-binding protein (MBP) from E. coli.
The protein is bound to a NeutrAvidin surface by a biotinylated
linker. MBP is labeled with a nonfluorescent acceptor
QSY7. The donor is Cy3.5, which is also bound to the surface
by a specialized linker (Figure 19.60). This linker contains
a cyclodextrin that binds to MBP as well as the Cy3.5
donor. These components are bound to the surface by a
biotinylated DNA linker arm that contains regions of single-
and double-stranded DNA. The single-stranded region
is present to allow changes in rigidity of the DNA by binding
of a complementary sequence called modulator DNA.
When both MBP and the linker are bound to the surface the
Figure 19.60. Linker arm containing the Cy3.5 donor and the cyclodextrin ligand. The fluorophore is bound to the surface via a biotinylated DNA
oligomer with a single stranded region for binding of modulator DNA. The lower panels show the change in donor intensity of the sensor (Figure
19.58) in response to maltose. Revised and reprinted with permission from [220]. Copyright © 2004, American Chemical Society.