¹⁴N Mines Pulsed-ENDOR of Proximal Histidine and Heme of Aquometmyoglobin and Fluormetmyoglobin

¹⁴N Mines Pulsed-ENDOR of Proximal Histidine and Heme of Aquometmyoglobin and Fluormetmyoglobin

Previous $^{19}F\;and\;^{1,2}H$ electron-nuclear double resonance (ENDOR) study of fluorometmyoglobin (MbF) in frozen-solution state provided sensitive tools sensing subtle structural changes of the heme that are not obtainable from X-ray. [Fann et al., J. Am. Chem. Soc. 1995, 117, 6019] Because of the intrinsic inhomogeneouse EPR line broadening effect of MbF in frozen-solution state, detection of the intrinsic inhomogeneouse EPR line broadening effect of MbF in frozen-solution state, detection of the electronic and geometrical changes of the heme ring itself and the proximal histidine by using $^{14}N$ CW ENDOR was interfered. In the present study, hyperfine-sensitive $^{14}N$ Mims ENDOR technique of pulsed-EPR was employed to probe the changes. With two different $\tau$ values of 128 and 196 ns, $^{14}N$ ENDOR signals of the heme and proximal histidine were completely resolved at $g'_{II}(=g_e=2)$. This study present that X-band $^{14}N$ Mims ENDOR sequence can sensitively detect the small changes of the spin densities and p orbital populations of the proximal and the heme nitrogens, caused by ligand and pH variation of the distal site.