Picosecond Thermometer in the Amide I Band of Myoglobin

The amide I and II bands in myoglobin show a heterogeneous temperature dependence, with bands at 6.17 and $6.43\mu \mathrm{m}$ which are more intense at low temperatures. The amide I band temperature dependence is on the long wavelength edge of the band, while the short wavelength side has almost no temperature dependence. We compare concepts of anharmonic solid-state crystal physics and chemical physics for the origins of these bands. We suggest that the long wavelength side is composed of those amino acids which hydrogen bond to the hydration shell of the protein, and that temperature dependent bands can be used to determine the time it takes vibrational energy to flow into the hydration shell. We determine that vibrational energy flow to the hydration shell from the amide I takes approximately 20 ps to occur.

Figure 1

(a) The infrared absorption spectrum of sperm whale myoglobin as a function of temperature from 5 to 300 K (blue line to red line). (b) The spectrum from 5.8 to $6.6\mu \mathrm{m}$.

Figure 2

(a) The infrared difference absorption spectrum (5 K base) of sperm whale myoglobin as a function of temperature from 10 to 280 K (blue line to red line), from 2.8 to $8.0\mu \mathrm{m}$. (b) The difference of the amide I and amide II regions of Mb (5 K base) as a function of temperature.

Figure 3

(a) Fits of the temperature dependence of the IR absorbance of the $6.17\mu \mathrm{m}$ line in Mb (solid diamonds). The fit of Eq. (1) to this area is shown as the solid line (red line), and the fit to Eq. (2) is given by the dashed line (green). (b) The temperature derivative $dA/dT$ in OD/K of the absorbance at 50 (blue dashed line) and 280 K (red solid line) between 5.9 and $6.6\mu \mathrm{m}$.

Figure 4

(a) The pump-probe response at $6.00\mu \mathrm{m}$ for a Mb sample. The dashed (blue) line is 50 K, and the solid (red) line is 280 K. (b) The pump-probe response at $6.17\mu \mathrm{m}$ for a Mb sample. The solid (red) line is the response at 280 K, and the dot-dashed (blue) line is the response at 50 K. Fits to the rise in the long time response are shown as a dashed (blue) line for 50 K, and long-short dashed (red) line for 280 K.

Figure 5

The pump-probe response at $6.120\mu \mathrm{m}$ for Mb at 300 K, long time scan. The inset shows the signal expanded on a vertical scale. The dashed line in the inset is a fit to an exponential decay with time constant $500\pm 100\mathrm{ps}$.