Time-resolved fluorescence and anisotropy decay of the tryptophan in adrenocorticotropin-(1-24)

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Abstract
The direct time-resolved fluorescence anisotropy of the single tryptophan residue in the polypeptide hormone ACTH-(1-24) and the fluorescence decay kinetics of this residue (Trp-9) are reported. Two rotational correlation times are observed. One, occurring on the subnanosecond time scale, reflects the rotation of the indole ring, and the other, which extends into the ns range, is dominated by the complex motions of the polypeptide chain. The fluorescence lifetimes of the single tryptophan in glucagon (Trp-25) and the 23-26 glucagon peptide were also measured. In all cases the fluorescence kinetics were satisfied by a double-exponential decay law. The fluorescence lifetimes of several tryptophan and indole derivatives and 2 tryptophan dipeptides were examined to interpret the kinetics. In close agreement with the findings of others the tryptophan zwitterion exhibits emission wavelength dependent double-exponential decay kinetics. At 320 nm .tau.1 = 3.2 ns and .tau.2 = 0.8 ns, with .alpha.1 = 0.7 and .alpha.2 = 0.3. Above 380 nm only the 3.2 ns component is observed. By contrast the neutral derivative N-acetyltryptophanamide has a single exponential decay of 3.0 ns. The multiexponential decay kinetics of the polypeptides are discussed in terms of flexibility of the polypeptide chain and neighboring side-chain interactions.