Date of Award


Degree Type


Degree Name

Doctor of Philosophy


The fluorescence of tryptophyl residues in proteins has been used as a probe of the protein environment for some time. However the reasons for the unique fluorescence properties of tryptophan in solution are still poorly understood.;Part I of this thesis is concerned with studying the photophysics of indole and tryptophan derivatives in solution, as a function of solvent, deuteration and temperature. The time-resolved fluorescence behaviour was studied to examine the reasons for the double exponential decay kinetics of some tryptophan derivatives. The quenching of the fluorescence of tryptophan by hydrogen and deuterium ions was modelled and the results indicated a change in the pK of the zwitterion-cation equilibrium in the excited state. It was found that excited state reorientation of the polar solvent shell was necessary for double exponential kinetics to be observed. Both the solvent and the charge accepting abilities of the substituents in tryptophan derivatives appear to play a role in the non-radiative rates, and in the case of 5-methoxytryptophan the nature of the substituents directly affects the energy of the emitting state. These results are discussed in terms of a general kinetic scheme, and charge transfer from indole to the solvent is considered as a possible mechanism for excited state solvent complex formation.;Part II is concerned with the binding interaction between fluorescein (Fl) and high affinity anti-fluorescein antibody (anti-Fl IgG). Charge transfer between a tryptophyl residue in the binding site and the bound Fl was considered as a possible binding interaction. Fl forms complexes, probably due to charge transfer, with tryptophan, tyrosine and methionine in solution. The complex with tryptophan has a relatively high association constant. The spectral changes resulting when Fl binds to anti-Fl IgD indicate some heterogeneity in the binding interaction. The fluorescence of the tryptophyl residues of the protein is quenched on binding. However, there is no direct evidence that charge transfer complexes between Fl and tryptophan are necessarily present in the binding sites of all high affinity anti-Fl IgG molecules.



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