A Förster-resonance-energy transfer-based method for fluorescence detection of the protein redox state.
A method for fluorescence detection of a protein's redox state based on resonance energy transfer from an attached fluorescence label to the prosthetic group of the redox protein is described and tested for proteins containing three types of prosthetic groups: a type-1 copper site (azurin, amicyanin, plastocyanin, and pseudoazurin), a heme group (cytochrome c550), and a flavin mononucleotide (flavodoxin). This method permits one to reliably distinguish between reduced and oxidized proteins and to perform potentiometric titrations at submicromolar concentrations.
Kuznetsova
Sofya
S
Leiden Institute of Chemistry-Gorlaeus Laboratories, Leiden University, The Netherlands.
Zauner
Gerhild
G
Schmauder
Ralf
R
Mayboroda
Oleg A
OA
Deelder
André M
AM
Aartsma
Thijs J
TJ
Canters
Gerard W
GW
D001400:Azurin / Q000737:chemistry; D002232:Carbocyanines / Q000737:chemistry; D003300:Copper / Q000737:chemistry; D003574:Cytochrome c Group / Q000737:chemistry; D005418:Flavodoxin / Q000737:chemistry; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry; D010084:Oxidation-Reduction; D011199:Potentiometry; D011506:Proteins / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods
D002232:Carbocyanines; D003574:Cytochrome c Group; D005418:Flavodoxin; D005456:Fluorescent Dyes; D011506:Proteins; C085321:cyanine dye 5; D001400:Azurin; D003300:Copper; C031732:cytochrome C-550