This title appears in the Scientific Report :
2003
Please use the identifier:
http://hdl.handle.net/2128/673 in citations.
Please use the identifier: http://dx.doi.org/10.1021/ja034040p in citations.
Measurement of submicrosecond intramolecular contact formation in peptides at the single-molecule level
Measurement of submicrosecond intramolecular contact formation in peptides at the single-molecule level
We describe a single-molecule-sensitive method to determine the rate of contact formation and dissociation between tryptophan and an oxazine derivative (MR121) on the basis of measurements of the photon distance distribution. Two short peptides (15 and 20 amino acids) derived from the transactivatio...
Saved in:
Personal Name(s): | Neuweiler, H. |
---|---|
Schulz, A. / Böhmer, M. / Enderlein, J. / Sauer, M. | |
Contributing Institute: |
Zelluläre Signalverarbeitung; IBI-1 |
Published in: | Journal of the American Chemical Society, 125 (2003) S. 5324 - 5330 |
Imprint: |
Washington, DC
American Chemical Society
2003
|
Physical Description: |
5324 - 5330 |
PubMed ID: |
12720444 |
DOI: |
10.1021/ja034040p |
Document Type: |
Journal Article |
Research Program: |
Neurowissenschaften |
Series Title: |
Journal of the American Chemical Society
125 |
Subject (ZB): | |
Link: |
Get full text OpenAccess |
Publikationsportal JuSER |
Please use the identifier: http://dx.doi.org/10.1021/ja034040p in citations.
We describe a single-molecule-sensitive method to determine the rate of contact formation and dissociation between tryptophan and an oxazine derivative (MR121) on the basis of measurements of the photon distance distribution. Two short peptides (15 and 20 amino acids) derived from the transactivation domain of the human oncoprotein p53 were investigated. With the fluorophore attached at the N-terminal end of the flexible peptides, fluorescence of the dye is efficiently quenched upon contact formation with a tryptophan residue. The mechanism responsible for the efficient fluorescence quenching observed in the complexes is assumed to be a photoinduced electron-transfer reaction occurring predominantly at van der Waals contact. Fluorescence fluctuations caused by intramolecular contact formation and dissociation were recorded using confocal fluorescence microscopy with two avalanche photodiodes and the time-correlated single-photon-counting technique, enabling a temporal resolution of 1.2 ns. Peptides containing a tryptophan residue at positions 9 and 8, respectively, show contact formation with rate constants of 1/120 and 1/152 ns(-1), respectively. Whereas the rate constants of contact formation most likely directly report on biopolymer chain mobility, the dissociation rate constants of 1/267 and 1/742 ns(-1), respectively, are significantly smaller and reflect strong hydrophobic interactions between the dye and tryptophan. Fluorescence experiments on point-mutated peptides where tryptophan is exchanged by phenylalanine show no fluorescence quenching. |