Fluorescence of histones H1. A tyrosinate-like fluorescence emission in ceratitis capitata H1 at neutral pH values.

We have examined the fluorescent properties of histones H1, and of some peptides derived from them, from calf thymus and from the fruit fly Ceratitis capitata. The fluorescent emission spectrum of folded histone H1 from C. capitata at neutral pH is characterized by a maximum at 303 nm and a shoulder at 340 nm. The overall quantum yields of fluorescence do not increase upon folding, although the fluorescence of the single tyrosyl residue of calf H1 is enhanced when the protein folds. As expected, the excitation maximum of calf H1 is shifted to longer wavelengths when the protein folds and its position does not depend upon the wavelength at which the fluorescence is observed. However, Ceratitis H1 exhibits two excitation maxima. The first corresponds to emission at 303 nm and it is slightly redshifted upon protein folding, whereas the second, which corresponds to emission at 340 nm, is displaced from 280 nm in the denatured protein to approximately 285 nm in the folded histone. This suggests that the two tyrosyl residues of the insect histone behave as independent fluorophores. The shoulder at 340 nm does not appear at pH 2, even when the protein is folded. Titration to neutral pH values results in the appearance of the shoulder, the process being characterized by a pK'a approximately equal to 3.7. The fluorescence spectrum of insect histone has been resolved into the contributions of the individual tyrosyl residues and the results suggest that the emission at 340 nm originates in a tyrosinate that may be formed in the excited state by proton transfer to the carboxylate anion of a glutamyl residue. The results obtained from these experiments have also aided in resolving the near-UV circular dichroism spectrum of insect histone Barbero, J.L., Franco, L., Montero, F., and Morán, F. (1980) Biochemistry 19, 4080-4087) into the individual contributions of the tyrosyl residues.

This work has been supported in part by Grant 4239/79 from the Comision Asesora de Investigacion Cientifica y Tecnica (Spain). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

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