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The initial structural evolution of carboxymyoglobin (MbCO) following photodissociation of CO is studied using optically heterodyne-detected (OHD) transient grating (TG) spectroscopy. This method provides detailed dynamical information on the electronic and structural states of the heme protein following photoexcitation. The phase anisotropy of MbCO is found to develop on subpicosecond to picosecond time scales and is much greater than can be attributed to the symmetry of the heme dipole transition. Control studies of carboxyprotoheme and deoxymyoglobin were used to identify the components due to protein structural relaxation and thermal relaxation, respectively. A geometric decomposition of the MbCO grating signals into contributions relative to the molecular axes provides evidence that the protein effectively changes its shape within 500 fs following ligand dissociation. These anisotropic mass displacements are a signature of functionally important motions since they imply a certain degree of directionality or mode selective coupling to the response. The anisotropic relaxation and observed dynamics provide further evidence that the low-frequency collective modes of proteins play an important role in transducing reaction forces into functions.

Analytic relations are developed to describe the coupling between experimentally observed optically heterodyne-detected (OHD) transient grating (TG) signals and the underlying molecular dynamics of heme proteins. The heterodyne detection was implemented using a diffractive optical element to generate the phase-matched beam pattern along with a reference beam for OHD. The phase stability between the reference and signal fields using this approach is shown to be excellent over several hours of signal collection. Under small signal conditions this leads to an enhancement in signal-to-noise of several orders of magnitude. The grating excitation provides an inherent acoustic reference that can be used to determine the absolute signal phase. This enables separation of the real and imaginary components to the nonlinear susceptibility as well as determination of both the absorption anisotropy and its real counterpart, the phase anisotropy. The relationship between the phase anisotropy and the observed OHD TG signals is expressed in terms of the complex molecular polarizability specific to heme proteins but can be readily extended to all other systems. Access to the material anisotropy provides a direct probe of mode selective coupling in proteins, i.e., the nonuniform displacements determined by the underlying asymmetric protein structure.

The initial structural evolution of carboxymyoglobin (MbCO) following photodissociation of CO is measured using a recently developed method of heterodyne detection. This method increases the sensitivity, generates tilted pulse fronts which exactly cancel temporal broadening in noncollinear beam geometries, and provides an inherent acoustic reference that enables unambiguous separation of the real and imaginary components to the nonlinear susceptibility. With the latter advance, both the absorption anisotropy and its real counterpart, the phase anisotropy, can be measured with this technique. Access to the real part of the material anisotropy provides new information that can be correlated to the bath dynamics. In the MbCO studies, the phase anisotropy is found to develop on picosecond time scales and is much greater than can be attributed to the symmetry of the heme dipole transition. This provides direct evidence that the shape of the protein changes in the first few picoseconds following photodissociation. The asymmetric nature of the protein structure presumably plays an integral role in the development of the nonuniform displacements. The magnitude of the phase anisotropy and observed dynamics give strong evidence that the low-frequency collective modes of the protein are involved in transducing the reaction forces of the Fe ligation site into directed motions associated with the initial oxy to deoxy tertiary structure change.