| A new fluorescent Fmoc-SPPS compatible amino acid that incorporates the halide-sensitive 6-methoxyquinolinium group was synthesized and incorporated in two peptides with known affinity for heparin. Sensor number one, AGf73, showed a 320% increase in fluorescence intensity upon heparin association, with an apparent dissociation constant of 2.6 muM at 100 mM NaCl. AGf73 selectivity was illustrated by the absence of changes in fluorescence intensity in the presence of chondroitin sulfate A. Furthermore, the proposed chloride-dependent sensing mechanism was supported by the fact that fluoride ions did not elicit a significant fluorescence change in the presence or in the absence of heparin. Sensor number two, MK2f-69d-61, was based on the second domain of human midkine, under the same working principle as AGf73, adding an internal fluorescent moiety that would render it ratiometric. Although adequate folding of the second sensor was not achieved, the ratiometric nature of the probe was corroborated experimentally.1.;1This work has been submitted to ChemBioChem and is being revised for publication. |
