Novel fluorescent sensing system for α-fructosyl amino acids based on engineered fructosyl amino acid binding protein

A novel fluorescent sensing system for a-glycated amino acids was created based on fructosyl amino acid binding protein (FABP) from Agrobacterium tumefaciens. The protein was found to bind specifically to the a-glycated amino acids fructosyl glutamine (Fru-Gln) and fructosyl valine (Fru-Val) while not binding to epsilon-fructosyl lysine. An Ile166Cys mutant of FABP was created by genetic engineering and modified with the environmentally sensitive fluorophore acrylodan. The acrylodan-conjugated mutant FABP showed eight-fold greater sensitivity to Fru-Val than the unconjugated protein and could detect concentrations as low as 17 nM, making it over 100-fold more sensitive than enzyme-based detection systems. Its high sensitivity and specificity for a-substituted fructosyl amino acids makes the new sensing system ideally suited for the measurement of hemoglobin Alc (HbAlc), a major marker of diabetes. (c) 2006 Published by Elsevier B.V.