Inhomogeneous broadening of the 760-nanometer photoproduct band of carboxymyoglobin at cryogenic temperatures has been demonstrated with a dynamic hole burning technique. Line-shape changes and frequency shifts in this spectral band are generated by ligand recombination and are shown not to be the result of structural relaxation below 60 K. The observation of dynamic hole burning exposes the relation between the structural disorder responsible for the inhomogeneous broadening and the well-known distributed ligand rebinding kinetics. The findings provide direct evidence for the functional relevance of conformational substates in myoglobin rebinding. In addition, a general protocol for evaluating the relative contributions of structural relaxation and hole burning to the spectral changes accompanying ligand rebinding in hemeproteins is presented.
ASJC Scopus subject areas