On the involvement of electron transfer reactions in the fluorescence decay kinetics heterogeneity of proteins

Abstract

Time-resolved fluorescence study of single tryptophan-containing proteins, nuclease, ribonuclease T1, protein G, glucagon, and mastoparan, has been carried out. Three different methods were used for the analysis of fluorescence decays: the iterative reconvolution method, as reviewed and developed in our laboratory, the maximum entropy method, and the recent method that we called "energy transfer" method. All the proteins show heterogeneous fluorescence kinetics (multiexponential decay). The origin of this heterogeneity is interpreted in terms of current theories of electron transfer process, which treat the electron transfer process as a radiationless transition. The theoretical electron transfer rate was calculated assuming the peptide bond carbonyl as the acceptor site. The good agreement between experimental and theoretical electron-transfer rates leads us to suggest that the electron-transfer process is the principal quenching mechanism of Trp fluorescence in proteins, resulting in heterogeneous fluorescence kinetics. Furthermore, the origin of apparent homogeneous fluorescence kinetics (monoexponential decay) in some proteins also can be explained on the basis of electron-transfer mechanism.

Variation of fluorescence in tissue with temperature

“…As tryptophan residue becomes hydrogen bonded or exposed to water its emission spectra and quantum yield is changed. There are several factors that determine the emission from tryptophan residues of protein molecules such as (1) quenching by proton transfer from nearby charged amino groups, (2) quenching by electron acceptors such as protonated carboxyl groups, (3) electron transfer quenching by disulfides and amides, (4) electron transfer quenching by peptide bonds in the protein backbone, and (5) resonance energy transfer among the tryptophan residues [16][17][18][19][20][21]. Additionally, a protein may exist in more than a single conformation, with each displaying a different quantum yield.…”

P2X3 receptor gating near normal body temperature

“…In the nervous system, ATP acts as an excitatory neurotransmitter in many regions of the CNS [7,[19][20][21][22][23]; in the periphery, ATP is known to activate nociceptive fibers and produce pain sensation [3,16]. Rapid excitatory effects of ATP are mediated through multigene family of P2X [1][2][3][4][5][6][7] receptors, which open the cation-selective ion-conducting pore upon agonist binding [15,17]. Purinoreceptors of P2X 3 subtype are expressed almost exclusively in mammalian sensory neurons and are thought to serve several specific functions from coding temperature sensation to nociception [18,29].…”

Fluorescence Decay Heterogeneity Model Based on Electron Transfer Processes in an Enzyme-Ligand Complex

“…electron transfer (and/or hole transfer) or fluorescence resonance energy transfer (FRET) from excited tryptophan and/or tyrosine residues. Since no indication for energy acceptors for FRET in proteins have been found [23], we consider fluorescence decay modeling which involves electron transport processes. The general idea is that the electron of an excited fluorophore can migrate (with a transfer rate ν(t)) to some excited states different from the initial one.…”

“…One example of such a process in proteins is electron transport to a neighboring quenching residue. Many possible electron acceptors have been proposed, such as a peptide bond carbonyl group [23] (due to appearance of a net positive charge on the peptide bond carbonyl carbon [24]) or amino acid residues involved in π−π stacking interaction. Since almost all amino acid side chains are more or less effective quenchers [10,25], the proposed model is applicable to proteins in general.…”