Probing redox proteins on a gold surface by single molecule fluorescence spectroscopy.
The interaction between the fluorescently labeled redox protein, azurin, and a thin gold film is characterized using single-molecule fluorescence intensity and lifetime measurements. Fluorescence quenching starts at distances below 2.3 nm from the gold surface. At shorter distances the quantum yield may decrease down to fourfold for direct attachment of the protein to bare gold. Outside of the quenching range, up to fivefold enhancement of the fluorescence is observed on average with increasing roughness of the gold layer. Fluorescence-detected redox activity of individual azurin molecules, with a lifetime switching ratio of 0.4, is demonstrated for the first time close to a gold surface.
Leiden Institute of Physics, Leiden University, Huygens Laboratory, Niels Bohrweg 2, 2333CA Leiden, The Netherlands.
Abdalmohsen T
AT
Elmalk
Jante M
JM
Salverda
Leandro C
LC
Tabares
Gerard W
GW
Canters
Thijs J
TJ
Aartsma
D001400:Azurin / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry*; D006046:Gold / Q000737:chemistry*; D055767:Immobilized Proteins / Q000737:chemistry*; D008958:Models, Molecular; D010084:Oxidation-Reduction; D011550:Pseudomonas aeruginosa / Q000737:chemistry*; D013050:Spectrometry, Fluorescence / Q000379:methods*; D013499:Surface Properties
D001426:Bacterial Proteins; D055767:Immobilized Proteins; D001400:Azurin; D006046:Gold