FLUORESCENCE OF TYROSINE AND TRYPTOPHAN IN PROTEINS USING ONE‐ AND TWO‐PHOTON EXCITATION

Abstract— We examined the emission spectra of tyrosine‐ and tryptophan‐containing proteins using one‐photon (270–310 nm) and two‐photon (565–610 nm) excitation. Emission spectra for two‐photon excitation of native and denatured human serum albumin and of three purine nucleoside phosphorylases indicated an absence of the tyrosine emission normally seen for one‐photon excitation below 290 nm. We examined the one‐photon and two‐photon excitation spectra of tyrosine‐tryptophan mixtures to determine the origin of selective excitation of the tryptophan residues. These results confirmed a short‐wavelength shift of the tyrosine two‐photon excitation spectrum relative to that of tryptophan, as recently reported by Rehms and Callis (1993) Chem. Phys. Lett. 208, 276–282.