Construction of a thiamin sensor from the periplasmic thiamin binding protein.
This communication describes the development of a thiamin sensor based on the bacterial thiamin binding protein. A triple mutant (C48S, C50S, S62C) of TbpA was labeled on C62 with N-[2-(L-maleimidyl)ethyl]-7-(diethylamino)coumarin-3-carboxamide (MDCC). Thiamin binding to this protein reduced the coumarin fluorescence giving a thiamin sensor with low nanomolar sensitivity.
Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY 14853, USA.
Jeremiah W
JW
Hanes
Debashree
D
Chatterjee
Erika V
EV
Soriano
Steven E
SE
Ealick
Tadhg P
TP
Begley
D019943:Amino Acid Substitution; D015374:Biosensing Techniques / Q000379:methods*; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003374:Coumarins / Q000737:chemistry; D007700:Kinetics; D009154:Mutation; D019897:Periplasm / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D013831:Thiamine / Q000032:analysis*
D002352:Carrier Proteins; D003374:Coumarins; C026989:thiamine-binding protein; C088410:N-(2-(1-maleimidyl)ethyl)-7-(diethylamino)coumarin-3-carboxamide; D013831:Thiamine