A state-of-the-art instrument for the measurement of fluorescence lifetime and related parameters by the phase-shift method is requested. In comparison to an existing outdated pulse-excitation instrument, the requested instrument is superior in its sensitivity, spectral and temporal resolution, optical design for polarized measurements and software for data analyses. The instrument will be used by a group of investigators who have been using fluorescence techniques to study various aspects of muscle contraction and its regulation and problems related to the properties of the lens of the eye. The projects centered on muscle include the mechanism whereby chemical energy stored in ATP is transduced into mechanical energy the mechanism whereby skeletal, cardiac and smooth muscles are regulated by the Ca2+ released into the sarcoplasm upon neural stimulation, and the mechanism whereby Ca2+ is returned into the sarcoplasmic reticulum during muscle relaxation; eye related work deals with changes occurring in components of eye tissues (vitreous. lens) upon photo-oxidation. The requested instrument will enhance our research effort in the following types of studies: 1) Fluorescence spectrum and lifetime measurements to define and monitor protein conformational states. 2) Resonance energy transfer measurements to map the proximity relationships between sites in each protein. 3) Anisotropy decay measurements to study the shape and segmental flexibility of each protein. 4) Solute quenching measurements to examine the mode of interactions between the proteins. Information derived from these and other approaches will provide an understanding at the molecular level of the functioning of the striated muscle of the skeletal system and the heart, as well as of smooth muscles in internal organs and the vascular system. Similarly, information about changes in macromolecular components of the eye will help in the elucidation of normal and pathological processes, particularly those induced by radiation.
