Tryptophan residue model

The electron transfer from the photoexcited Ru(bpy)32+ to MV2+ confined in a polysiloxane film showed a complete static quenching following the Perrin model,32) and the electron transfer distance rc was 1.4 nm, which is comparable to a conventional electron transfer distance in biological systems. The presence of a tryptophan residue model, 3-methyindole (IND) enhanced much the quenching efficiency (Fig. 19.6) by lengthening the electron transfer distance, and the electron transfer distance was estimated to be 2.7 nm, almost twice that without the mediator.32 ... 

The most typical case is probably that in which tryptophan residues undergo rotations during the excited-state lifetime with a small angular amplitude (up to 30°).(30 69) Similar motions are observed in protein-dye complexes. We will consider below the qualitative models which have been put forward to describe such motions. 

Considerable red-edge effects exhibiting a dependence on the viscosity of the medium are observed for model solutions of indole and tryptophan(33) (Figure 2.11a), which permits this approach to be applied to studies of the dynamics of the environment of tryptophan residues in proteins. In discussion... 

The anisotropy decay of the tryptophan fluorescence of both model peptides and biologically active peptides containing a single tryptophan residue has been determined in various studies. Even in the case of the tripeptide H-Gly-Trp-Gly-OH quenched by acrylamide the anisotropy decay displayed two correlation times with values of 39 and 135 ps. 44 The shorter correlation time was thought to be due to motions of the indole ring relative to the tripeptide. In the case of ACTH(l-24) the fluorescence anisotropy decay of the single tryptophan residue in position 9 of the peptide sequence obtained in phosphate buffer (pH 7, 3.5 °C) was also double-exponential. 29 The shorter rotational correlation time (0 = 92ps)... 

The chemical synthesis of the Amanita toxins has presented several problems, in particular those related to the formation of the sulfur bridge. The latter has been explored with model compounds.[2 31 It has been found that the synthesis of the (sulfanyl)indole moiety can be achieved by reacting an indole compound with an alkanesulfenyl chloride. A model tryptathionine compound has been prepared by reacting A-acyl-L-cysteine and /V-acyl-L-tryptophan in the presence of A-chlorosuccinimide in glacial acetic acid at room temperature.[4] The sulfanylation reaction has been subsequently exploited for the selective chemical modification of tryptophan residues in proteins using 2-nitrophenylsulfenyl chlorideJ5 ... 

All lipases of this family are characterized by a helical lid that covers the active site (Kg. 9X It has been hypothesized [145] that this lid moves away in a hydrophobic environment, thereby making the active site accessible to the lipid substrate. In this hypothesis an important role has been reserved for the bulky tryptophane residue at position 117 in the center of the helical lid. In the Rhizoptts javwicus lipase, however, this bulky hydrophobic residue is replaced by a alanine, and no other bulky residue seems located so that it could take die role of residue 117. Therefore, it was concluded that the lipid-interaction model that haa been suggested for this class of lipase is not generally applicable in all its details. 

Pro-oxidant conditions are favoured in infant formulae by the presence of iron and of vitamin C and can lead to oxidative damage to tryptophan residues, which here is of particular importance, tryptophan often being the limiting amino acid. Using a-lactalbumin as a model compound, as it is high in tryptophan, Puscasu and Birlouez-Aragon484 studied the loss of fluorescence due to tryptophan (Aex=290/Aem=340 nm) on incubation with lactose, preformed early and advanced MRP (from proteose-peptone, because it is low in tryptophan), H202/Fe2+, or ascorbate/Fe3+. In each case, after 3 h, there was an appreciable loss of tryptophan from the pH 4.6-soluble protein of about 28%. The MRPs, both formed and preformed, exhibited fluorescence at Aex = 350/Aem = 435-440 (major) and Aex = 330/Aem = 420 nm. 

In his pioneering contribution, Porath postulated that the histidine, cysteine, and tryptophan residues of a protein were most likely to form stable coordination bonds with chelated metal ions at near neutral pH [2]. To date, an analysis of several protein models [7] lend full to his original theory. Having said that, histidine, by far and away, plays the most prominent role in IMAC binding. In a very real sense, IMAC has subtly become synonymous as a histidine affinity technique. The absence of a histidine residue on a protein surface correlates with the lack of retention of that protein on any IDA-metal column. The presence of even a single histidine on a protein surface, available for coordination, results in retention of that protein on an IDA-Cu(II) column. Also, a protein needs to display at least two histidine residues on its surface to be retained on an IDA-Ni(II) column. Thus, beyond its role as a purification technique, IMAC has been used as a tool to probe the surface topography of proteins [6]. 

Fig. 6. Organization of apolipoprotein molecules in discoidal HDL particles. (A) Double belt model for apo A1 structure at the edge of discoidal HDL complex. Two ring-shaped molecules of apo A1 are stacked on top of each other with both molecules in an anti-parallel orientation, allowing the helix registry to maximize intermolecular salt-bridge interactions. Only the charged residues at selected positions are explicitly displayed. (B) Model of apo E in discoidal HDL complex depicting the locations of engineered tryptophan residues on helix 4. Fluorescence from these amino acids was monitored to determine helix orientation. Two out of a total of about four mole-cules/particles of apo E are depicted in which the helical axes are oriented perpendicular to the PL acyl chains.

Fluorescence intensity decay of the Trp residues ( ex, 300 nm) yields four fluorescence lifetimes, 0.22, 0.44, 2.06 and 4.46 ns. The authors considered the two shortest lifetimes as the result of the excited state quenching of the tryptophan emission by the protein backbone, while the two longest lifetimes are characteristics of the emission of the Trp residues from two different environments (Kung et al. 1998). Tryptophan free in solution emits with two lifetimes, 0.5 and 3.1 ns. In this case, the short lifetime cannot be assigned to the presence of a backbone protein. The two lifetimes are considered to be the result of the different side chain retainers conformations. Thus, a tryptophan in presence of different environments, water and protein matrix, can have the same or identical fluorescence lifetimes. This example shows clearly that it is not obvious to explain the origin of the fluorescence lifetime of tryptophan residues in proteins by the rotamers model. 

Fluorescence emission lifetime of one tryptophan residue located in five different positions within an 18-residue amphiphatic peptide (Table 7.5) was also analyzed with the rotamers model. These peptides are unstructured in aqueous solution and become stmctured when associated to a lipid. A blue shift in the emission maximum of the Trp residue is observed, indicating that the fluorophore is in contact with a hydrophobic environment when the lipid is bound to the peptides. Only peptide 18D-12 shows an emission maximum located at 354 revealing that in this peptide tryptophan is in contact with aqueous environment. These data are consistent witli the fact that association of lipid with peptide yield an a-helix where the tryptophan residues located on the hydrophobic surface of the a-helix are in the hydrophobic environment provided by the lipid surface, whereas tryptophans located on the hydrophilic face of the helix are directed toward the aqueous phase. Also, the helix axis of the peptides is parallel to the lipid surface (Fig. 7.9). 

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