STRUCTURAL FACTORS CONTROLLING LIGAND-BINDING TO MYOGLOBIN - A KINETIC HOLE-BURNING STUDY

Using temperature-derivative spectroscopy in the temperature range bel ow 100 K, we have studied the dependence of the Soret band on the reco mbination barrier in sperm whale carbonmonoxy myoglobin (MbCO) after p hotodissociation at 12 K. The spectra were separated into contribution s from the photodissociated species, MbCO, and CO-bound myoglobin, Th e line shapes of the Soret bands of both photolyzed and liganded myogl obin were analyzed with a model that takes into account the homogeneou s bandwidth, coupling of the electronic transition to vibrational mode s, and static conformational heterogeneity. The analysis yields correl ations between the activation enthalpy for rebinding and the model par ameters that characterize the homogeneous subensembles within the conf ormationally heterogeneous ensemble. Such couplings between spectral a nd functional parameters arise when they both originate from a common structural coordinate. This effect is frequently denoted as ''kinetic hole burning.'' The study of these correlations gives direct insights into the structure-function relationship in proteins. On the basis of earlier work that assigned spectral parameters to geometric properties of the heme, the connections with the heme geometry are discussed. We show that two separate structural coordinates influence the Soret lin e shape, but only one of the two is coupled to the enthalpy barrier fo r rebinding, We give evidence that this coordinate, contrary to widesp read belief, is not the iron displacement from the mean heme plane.