Show simple item record

dc.contributor.authorRyder, Alan G.
dc.contributor.authorSzczupak, Boguslaw
dc.contributor.authorTogashi, Denisio M.
dc.contributor.authorCalvet, Amandine
dc.contributor.authorO'Loughlin, Muireann
dc.date.accessioned2014-01-21T15:10:18Z
dc.date.available2014-01-21T15:10:18Z
dc.date.issued2009
dc.identifier.citationTogashi, DM,Szczupak, B,Ryder, AG,Calvet, A,O'Loughlin, M (2009) 'Investigating Tryptophan Quenching of Fluorescein Fluorescence under Protolytic Equilibrium'. Journal Of Physical Chemistry A, 113 :2757-2767.en_US
dc.identifier.urihttp://hdl.handle.net/10379/3984
dc.description.abstractFluorescein is one of most used fluorescent labels for characterizing biological systems, such as proteins, and is used in fluorescence microscopy. However, if fluorescein is to be used for quantitative measurements involving proteins then one Must account for the fact that the fluorescence of fluorescein-labeled protein can be affected by the presence of intrinsic amino acids residues, such as tryptophan (Trp). There is a lack of quantitative information to explain in detail the specific processes that are involved, and this makes it difficult to evaluate quantitatively the photophysics of fluorescein-labeled proteins. To address this, we have explored the fluorescence of fluorescein in buffered solutions, in different acidic and basic conditions, and at varied concentrations of tryptophan derivatives, using steady-state absorption and fluorescence spectroscopy, combined with fluorescence lifetime measurements. Stern-Volmer analyses show the presence of static and dynamic quenching processes between fluorescein and tryptophan derivatives. Nonfluorescent complexes with low association constants (5.0-24.1 M-1) are observed at all pH values studied. At low pH values, however, an additional static quenching contribution by a sphere-of-action (SOA) mechanism was found. The possibility of a proton transfer mechanism being involved in the SOA static quenching, at low pH, is discussed based on the presence of the different fluorescein prototropic species. For the dynamic quenching process, the bimolecular rate constants obtained (2.5-5.3 x 10(9) M(-1)s(-1)) were close to the Debye-Smoluchowski diffusion rate constants. In the encounter controlled reaction mechanism, a photoinduced electron transfer process was applied using the reduction potentials and charges of the fluorophore and quencher, in addition to the ionic strength of the environment. The electron transfer rate constants (2.3-6.7 x 10(9) s(-1)) and the electronic coupling values (5.7-25.1 cm (1)) for fluorescein fluorescence quenching by tryptophan derivatives in the encounter complex were then obtained and analyzed.en_US
dc.formatapplication/pdfen_US
dc.language.isoenen_US
dc.relation.ispartofJournal Of Physical Chemistry Aen
dc.subjectPhotoinduced electron-transferen_US
dc.subjectBovine serum-albuminen_US
dc.subjectSteady-stateen_US
dc.subjectProton-transferen_US
dc.subjectRate constantsen_US
dc.subjectOrganic-dyesen_US
dc.subjectProteinen_US
dc.subjectMoleculesen_US
dc.subjectLifetimeen_US
dc.subjectSpectroscopyen_US
dc.titleInvestigating tryptophan quenching of fluorescein fluorescence under protolytic equilibriumen_US
dc.typeArticleen_US
dc.date.updated2013-10-25T09:29:35Z
dc.identifier.doihttp://dx.doi.org/10.1021/jp808121y
dc.local.publishedsourcehttp://dx.doi.org/10.1021/jp808121yen_US
dc.local.publisherstatement"Reprinted (adapted) with permission from Investigating trypthopan quenching of fluorescein fluorescence under protolytic equilibrium. D.M. Togashi, B. Szczupak, A.G. Ryder, A. Calvet, and M. O¿Loughlin, Journal of Physical Chemistry A, 113(12), 2757-2767, (2009). Copyright 2009 American Chemical Society."en_US
dc.description.peer-reviewedpeer-reviewed
dc.contributor.funder|~|
dc.internal.rssid1339315
dc.local.contactAlan Ryder, School Of Chemistry, Room 228, Arts/Science Building, Nui Galway. 2943 Email: alan.ryder@nuigalway.ie
dc.local.copyrightcheckedNo
dc.local.versionACCEPTED
nui.item.downloads883


Files in this item

Attribution-NonCommercial-NoDerivs 3.0 Ireland
This item is available under the Attribution-NonCommercial-NoDerivs 3.0 Ireland. No item may be reproduced for commercial purposes. Please refer to the publisher's URL where this is made available, or to notes contained in the item itself. Other terms may apply.

The following license files are associated with this item:

Thumbnail

This item appears in the following Collection(s)

Show simple item record