Resonance Raman spectroscopic study of the neutral flavin radical complex of DNA photolyase from Escherichia coli

The neutral Ravin radical complex of DNA photolyase from Escherichia coli w as studied by resonance Raman spectroscopy for the first time. The experime nts were carried out with enzyme variants that lack the light-harvesting me thenyltetrahydrofolate cofactor so that the resonance Raman spectra exclusi vely display the vibrational bands of the Ravin radical in the catalytic si te. Spectral changes induced upon H-D exchange and upon substituting the na tural flavin by its N-15-labelled isotopomer allow a tentative assignment o f the prominent bands in the region between 1200 and 1650 cm(-1). It is sho wn that most of the bands observed with 568 run excitation originate from m odes localized in the isoalloxazine ring. In conjunction with previous data on flavin radicals, the present results allow the identification of the ma rker bands that appear to be particularly sensitive towards changes in the electron density distribution of this ring and, hence, reflect the specific intermolecular interactions in the catalytic site. Copyright (C) 2001 John Wiley & Sons, Ltd.