Assignment of the heterogeneous static and time-resolved tryptophan fluorescence of 3-phosphoglycerate kinase

The heterogeneous fluorescence of yeast 3-phosphoglycerate kinase, a hinge-bending enzyme with two tryptophan residues, has been resolved into three emission components using steady-state and time-resolved studies of the fluorescence quenching by acrylamide, iodide and caesium ions at different emission wavelengths. The buried Trp-333 has a blue-shifted heterogeneous emission spectrum characterised by three fluorescence lifetimes, and is inaccessible to quenchers. The surface Trp-308 also has a heterogeneous emission with multiple lifetimes. The emission of Trp-308 can be separated into a blue-shifted emission accessible to acrylamide and caesium only, and a red-shifted emission accessible to all three quenchers.