5-HYDROXYTRYPTOPHAN - AN ABSORPTION AND FLUORESCENCE PROBE WHICH IS ACONSERVATIVE REPLACEMENT FOR [A14 TYROSINE] IN INSULIN

Use of insulin's intrinsic tyrosine absorption and fluorescence to mon itor its interaction with the insulin receptor is limited because the spectral properties of the receptor tryptophan residues mask the spect ral properties of the hormone tyrosine residues. We describe the synth esis of an insulin analog where A14 tyrosine is replaced by a tryptoph an analog, 5-hydroxytryptophan. This insulin is spectrally enhanced si nce 5-hydroxytryptophan has an absorption band above 300nm which is at lower energies than the absorption of tryptophan. Steady-state and ti me-resolved fluorescence parameters indicate that 5-hydroxytryptophan reports the same information about the environment of the A14 side cha in as does the corresponding tryptophan-containing insulin. The synthe tic hormone is a full agonist compared to porcine insulin, but has sli ghtly reduced specific activity. Consequently, this spectrally enhance d insulin analog will be useful for hormone-receptor interaction studi es since it can be observed by both absorption and fluorescence even i n the presence of the tryptophan-containing receptor.