Observation of deuterium-labeled diacetyldeuteroporphyrin incorporated in cyanoferrimyoglobin by deuterium nuclear magnetic resonance.

Sperm whale apomyoglobin was reconstituted with selectively deuterated D₆-2,4-diacetyldeuterohemin in which the ²H label was confined to the methyl groups of the acetyl moieties. A single resonance was observed in ²H NMR of the cyanoferrimyoglobin derivative, with a chemical shift 0.80 ppm downfield of external D₁₂-TMS at pH 6.7. The corresponding chemical shift of D₆-2,4-diacetyldeuterohemin-OMe as the cyanide complex in pyridine-water was 0.96 ppm downfield of external D₁₂-TMS. The prominent HOD peak was well separated at 4.4 ppm downfield. The line width of the porphyrin ²H resonances in both the protein and free solvent environments yields evidence of considerable rotational freedom of the -CD3 groups about their axes.