Novel fluorescent sensing system for alpha-fructosyl amino acids based on engineered fructosyl amino acid binding protein.
A novel fluorescent sensing system for alpha-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 alpha-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 alpha-substituted fructosyl amino acids makes the new sensing system ideally suited for the measurement of hemoglobin A1c (HbA1c), a major marker of diabetes.
Sakaguchi
Akane
A
Department of Biotechnology, Tokyo University of Agriculture and Technology, Koganei, Tokyo 184-8588, Japan.
Ferri
Stefano
S
Tsugawa
Wakako
W
Sode
Koji
K
D000596:Amino Acids / Q000032:analysis* / Q000737:chemistry; D015374:Biosensing Techniques; D002352:Carrier Proteins; D005453:Fluorescence; D005632:Fructose / Q000737:chemistry*; D015202:Protein Engineering; D013379:Substrate Specificity
D000596:Amino Acids; D002352:Carrier Proteins; D005632:Fructose