Resonance Raman Studies of Electron-Nuclear Coupling, Time Resolved Dynamics, and Magnetic Perturbations of Biomolecules

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The objectives of this research are to achieve a better understanding of the optical properties of biological molecules, both in and out of equilibrium, and to apply this knowledge to elucidate their mechanism of action. Resonance Raman scattering and its time domain analog, femtosecond coherence spectroscopy (FCS) will be used to probe the electronic and nuclear structure/function relationships in heme proteins like myoglobin, hemoglobin and cytochrome c. Related techniques such as electronic absorption, pump-probe kinetics, fluorescence, infrared, nuclear resonance vibrational (NRVS) spectroscopy, and magnetic field perturbations will also be used to characterize samples and aid in the assignment of the low frequency motions revealed using novel FCS techniques developed in the prior award period. Picosecond methodology will be developed using synchronized self-modelocked Ti:sapphire lasers to carry out both time resolved resonance Raman scattering and to extend the range of kinetic studies into the ns regime. Dynamic absorption lineshapes will be measured using broad-band continuum techniques so that time dependent resonance conditions can be properly incorporated into the analysis of vibrational relaxation obtained from the Stokes and anti-Stokes Raman scattering. Theoretical advances in understanding the absolute magnitude and phase of the resonant FCS signals will be exploited to eliminate non-oscillatory background signals and allow improved detection of low frequency modes. Phase and amplitude excitation profiles will be carried out to reveal inhomogeneities and anharmonicities associated with the low frequency motions. Low temperature measurement techniques will be developed so that FCS experiments can be carried out as a function of the sample "glass" transition. This project has a broad impact that underlies the field of protein dynamics that is crucial to our basic understanding of living systems. This work involves the development of state-of-the-art laser based spectroscopy that is applied to biomolecular systems. Instrumentation and theoretical development will enhance the infrastructure for doing time-resolved protein dynamics research by producing an effective FCS system that expands the use of this experimental technique by making it routinely accessible. These studies are significant not only in their relationship to the biological sciences, but also in their connection to our understanding of basic light-matter interaction associated with complex systems in the condensed phase. Graduate and undergraduate students will be trained in cutting edge optical and biological techniques that benefit society through the enhancement of the human resource infrastructure. This project is supported by the Molecular Biophysics Program in the Division of Molecular and Cellular Biosciences in the Directorate for Biological Sciences and the Division of Physics in the Mathematical and Physical Sciences Directorate.