Modication of Tryptophan
Tryptophan is responsible for most of characteristic protein UV absorbance at 280 nm; tryptophan is also responsible for most of the intrinsic uorescence of proteins. Tryptophan is also a target for photooxidation mediated by sensitizers such as methylene blue. There is further discussion of photooxidation in Chapter 5. For the purpose of the current discussion, photolysis is dened as the decomposition of substances on exposure to light, while photooxidation is the oxidation of a substance on exposure to light.24 Irradiation of tryptophan with UV light results in the formation of tryptophan radical and a hydrated electron25-30; the formation of hydrogen atoms from the photolysis of tryptophan has been reported.6,31 While the photolysis of tryptophan is still poorly understood, a mechanism where a hydrogen is removed from the indole nitrogen followed by rearrangement to yield a radical at C8 or C9 is consistent with the observations of Léonard and coworkers.32 One of the practical consequences of the photolysis of tryptophan is the action of tryptophan as a photosensitizer,33 which can modify other amino residues12,34 that contributed to the oxidation of a methionine residue in a monoclonal antibody.12 This oxidation is mediated by tryptophan radical-mediated formation of reactive oxygen species35 by the tryptophan radical, which forms hydrogen peroxide.36 Earlier work by Sidorkina and coworkers37 reported on the modication of E. coli Fpg protein by UVB light (280-315 nm) in 50 mM sodium phosphate, pH 7.4, containing 2 mM 2-mercaptoethanol and 10% glycerol which resulted in polymerization of the protein. Polymerization of the protein was observed as a result of UVB irradiation. Three of the ve tryptophanyl residues were modied in an air-saturated solution, while only one tryptophanyl residue was modied in an argon-saturated solution. Peptide bond cleavage is observed with the appearance of a fragment with a structure consistent with that of the rst 32 or 33 amino acids in the N-terminal region. The peptide bond cleavage is observed within either the air-saturated medium or the argonsaturated medium. It is suggested the cleavage results from photolysis of Trp34. Subsequent work
FIGURE 12.1 The structure of tryptophan and some related indole-derived products. Shown at the bottom is the photolytic reaction of tryptophan with chloroform to yield a formyltryptophan derivative (one of several possible isomers is shown). The reaction initially involved the formation of a tryptophan radical decaying with expulsion of a solvated electron that reacts with chloroform to yield a free radical that reacts with tryptophan decaying to yield the formyl derivative (Edwards, R.A., Jickling, G., and Turner, R.J., The light-induced reactions of tryptophan with halocompounds, Photochem. Photobiol. 75, 362-368, 2002).