SPECTROSCOPIC EFFECTS OF POLARITY AND HYDRATION IN THE DISTAL HEME POCKET OF DEOXYMYOGLOBIN

Distal pocket mutations at the E7 position (His64) of sperm whale deox ymyoglobin (deoxyMb) are used as a probe of distal pocket polarity and hydration, Changes of two key spectroscopic markers, the Fe-His(F8) s tretch in the resonance Raman spectrum and the position of band III in the absorption spectrum, are monitored as the His64Tyr, His64Phe, His 64Leu, and His64Gly mutations alter the distal heme pocket environment . The Fe-His vibration for the Phe, Leu, and Gly mutants is shifted to a lower frequency by 1-2 cm(-1) relative to the Tyr mutant, wild type (WT), and native deoxyMb, Band III shifts to the red by similar to 4 nm (similar to 70 cm(-1)) relative to WT and native deoxyMb for all th e His64 mutants examined in this work, We correlate the small shift in the Fe-His frequency to the local electrostatic environment directly above the heme iron. affected by the presence of a localized water mol ecule in the heme pocket that is hydrogen-bonded to the E7 residue. Th e position of band III is roughly correlated to the displacement of th e iron from the heme plane; however, the relatively large scatter in t his correlation, as well as its dependence on distal pocket mutations, suggests that the heme pocket environment, particularly the E7 residu e, also affects the energy of this transition.