Resonance Raman and FTIR spectra are reported for the oxidized azide-bound derivative of the quinol cytochrome bo(3) oxidase from Escherichia coli. The resonance Raman spectra display three isotope-dependent vibrational modes at 419, 2040, and 2061 cm(-1). The FTIR spectra display two isotope-dependent bands at 2040 and 2061 cm(-1). We assign the band at 419 cm(-1) to v(Fe-N-3-Cu-B) and the bands at 2040 and 2061 cm(-1) to v(as)(N-3) The observation of two v(as)(N-3) modes suggests that the azide ion binds to two different enzyme conformations, both forming bridging complexes with the binuclear center. Comparison of the FTIR data of the azide-bound cytochrome bo(3) and cytochrome aa(3) complexes reveal that there are quantitative differences in the structure of the heme o(3)-Cu-B and heme a(3)-Cu-B binuclear pockets upon azide binding. The present data on the vibrational frequencies of the azide-bound cytochrome bo(3) complex do not support the recent proposal that azide binds as a terminal ligand to CUB (Little, R. H.; Cheesman, M. R.; Thomson, A. J.; Greenwood, C.; Watmough, N. J. Biochemistry 1996, 35, 13780-13787) but an more reasonably interpreted to conclude that azide functions as a bridge between heme o(3) and Cu-B.