Geminate rebinding dynamics of nitric oxide to ferric hemoglobin in D2O solution.

Femtosecond mid-infrared (mid-IR) spectroscopy was used to probe geminate rebinding (GR) dynamics of photo-released nitric oxide (NO) to ferric hemoglobin (Hb(III)) in D2O solution at room temperature. Time-resolved vibrational spectra exhibit two overlapping NO bands for NO-bound Hb(III) (Hb(III)NO), a major band at 1925 cm(-1) (89%) and a minor one at 1905 cm(-1) (11%), suggesting that Hb(III)NO has at least two conformational substates. Both bands decay nonexponentially, each with a different time scale, and the decays are described by a stretched exponential function; the major band's decay is described by 0.96 exp(-t/40 ps)(0.86) + 0.04 and the minor band's decay is described by exp(-t/85 ps)(0.75). These decays arise mainly from the GR of the photo-released NO to Hb(III), indicating that the bound state's conformer influences the NO binding. In particular, the His64 residue, known to have inward conformation in the major band and outward conformation in the minor band, plays a significant role in controlling the binding of NO to Hb(III). The GR of NO to ferric Hb is slower than that to ferrous Hb, which shows fast and efficient GR due to the high reactivity of NO to the heme Fe(ii). The slower GR of NO to Hb(III) may be caused by the lower reactivity of NO to the heme Fe(iii).