Photoexcitation dynamics of azide ion bound ferric myoglobin probed by femtosecond infrared spectroscopy

The photoexcitation dynamics of N₃⁻-bound ferric myoglobin (MbN₃) were investigated after exciting MbN₃ in D₂O at 283 K with a 575 nm pulse by probing the anti-symmetric stretching mode of the azide. Global fitting of the overall time-resolved spectra revealed that thermal relaxation of two stretching bands proceeded with a time constant of 6 ps, and that a new absorption band formed and decayed with time constants of 0.6 and 23 ps, respectively. The new absorption near 2040 cm⁻¹ was attributed to the high-spin species 2.4 kJ/mol above the low-spin species, as the excited low-spin relaxes thermally via the high-spin species. However, this absorption could also arise from deligated N₃̄ remaining within the protein. The decay of this absorption can be interpreted as either spin transition of the high-spin species into the low-spin species or geminate recombination of the dissociated N₃̄. The implications of both interpretations are discussed.