Energy transfer to a proton-transfer fluorescence probe: Tryptophan to a flavonol in human serum albumin

Abstract

A protein f luorescence probe system, coupling excited-state intermolecular Förster energy transfer and intramolecular proton transfer (PT), is presented. As an energy donor for this system, we used tryptophan, which transfers its excitation energy to 3-hydroxyf lavone (3-HF) as a f lavonol prototype, an acceptor exhibiting excited-state intramolecular PT. We demonstrate such a coupling in human serum albumin-3-HF complexes, excited via the single intrinsic tryptophan (Trp-214). Besides the PT tautomer f luorescence ( max ‫؍‬ 526 nm), these protein-probe complexes exhibit a 3-HF anion emission ( max ‫؍‬ 500 nm). Analysis of spectroscopic data leads to the conclusion that two binding sites are involved in the human serum albumin-3-HF interaction. The 3-HF molecule bound in the higher affinity binding site, located in the IIIA subdomain, has the association constant (k 1 ) of 7.2 ؋ 10 5 M ؊1 and predominantly exists as an anion. The lower affinity site (k 2 ‫؍‬ 2.5 ؋ 10 5 M ؊1 ), situated in the IIA subdomain, is occupied by the neutral form of 3-HF (normal tautomer). Since Trp-214 is situated in the immediate vicinity of the 3-HF normal tautomer bound in the IIA subdomain, the intermolecular energy transfer for this donor͞ acceptor pair has a 100% efficiency and is followed by the PT tautomer f luorescence. Intermolecular energy transfer from the Trp-214 to the 3-HF anion bound in the IIIA subdomain is less efficient and has the rate of 1.61 ؋ 10 8 s ؊1 , thus giving for the donor͞acceptor distance a value of 25.5 Å.Excitation by light may initiate both intramolecular and intermolecular transformations of a molecule. Among excited-state intramolecular processes, the most important are electron transfer (1-4), proton transfer (PT; refs. 5 and 6), and conformational transformation (e.g., twisting, torsion, and isomerization; refs. 7-9). The most conspicuous intermolecular events are electron, proton, and energy transfer (10, 11). Excited-state transformations can be coupled with each other and with the surrounding medium. Dependence of the excitedstate intramolecular electron transfer on environmental relaxation (2, 3) and the twisting of solute groups in response to the excited-state charge transfer (12-15) are well-established phenomena. When joined in one molecular system, excited-state events can compete dynamically. An example of such competition is the mutual exclusion of intramolecular electron and PT in 4Ј-diethylamino-3-hydroxyflavone (16-19). Proper pairing of excited-state transformations can be a source of structural͞dynamical information about the molecules possessing these transformations, the excited-state transformations themselves, and the medium where the coupling takes place.The present paper focuses on the coupling between the Förster intermolecular energy transfer (11,20,21) and intramolecular PT (refs. 6 and 22; Fig. 1). The first step of such a coupling is the R Ϫ6 -dependent (R is the intermolecular distance) dipole-dipole intermolecular energy transfer from excited donor to unexcite...

A coupled proton-transfer and twisting-motion fluorescence probe for lipid bilayers

“…These changes generally are observed by monitoring the variation of photophysical properties, such as fluorescence anisotropy and lifetime, spectral position, and fluorescence quantum yield of the dye within the bilayers (1)(2)(3)(4). Recently, molecules showing excited-state protontransfer (ESPT) reactions have been suggested as probes for the study of protein conformation and binding-sites (5)(6)(7)(8). ESPT reactions are very fast elementary processes (9)(10)(11) involved in various area of physics, chemistry, and biology (6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18).…”

“…Recently, molecules showing excited-state protontransfer (ESPT) reactions have been suggested as probes for the study of protein conformation and binding-sites (5)(6)(7)(8). ESPT reactions are very fast elementary processes (9)(10)(11) involved in various area of physics, chemistry, and biology (6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18). Because of the observed large Stokes-shifted fluorescence of the proton-transfer band in these systems, such molecules might show some advantages with respect to others having a normal Stokes-shifted emission band.…”

Relaxation of the excited N-(2-hydroxy benzylidene) aniline molecule: An ab initio and TD DFT study

“…Understanding of the ESIPT process has invoked colossal interest among the photo physicists because of its applications in the development of laser dyes, 5 photo stabilizers 6 and scintillation counters. 7 Moreover, ESIPT reactions are used as optical probes for biomolecules 8 and in the investigation of biological environment. 9 The ESIPT reaction occurs in the molecular systems where the closed proximate proton donor and acceptor groups are connected in general by a six-member hydrogen bond network.…”

Síntesis de Derivados Fotoactivos de Ácidos Biliares para Aplicaciones Biológicas

“…Tal como se había mencionado en la introducción de este capítulo, la HSA posee tan sólo un triptófano (Trp), ubicado en el sitio 1, y la emisión de HSA puede ser sensible a la presencia de un aceptor de energía. [137] En este contexto, se estudió la fluorescencia de HSA (que corresponde a la emisión de Trp con un máximo a 338 nm, λ exc 290 nm) tras adición de cantidades crecientes de los derivados de NBD.…”

“…1, p. 43), tomando para Trp τ 0 = 6.38 ns, [137] se obtuvieron las correspondientes constantes de velocidad de desactivación que fueron de k q > 10 12 M -1 s -1 (Figura 5.8b para dos ejemplos), mucho mayor que la constante de difusión en biopolímeros, [138] indicando así el carácter estático de esta desactivación. En este caso, los datos se analizaron también con una ecuación de Stern-Volmer modificada: [139] (Ec.…”