This is a proposal to acquire a stopped flow circular dichroism spectrometer. The instrument will be used to monitor circular dichroism of samples of biological macromolecules on a stopped flow (millisecond) time scale. The primary use of the instrument at Texas A&M University will be to monitor protein rcfolding. There are currently 6 research groups studying various aspects of protein folding, and all have immediate need of the instrument. In addition, there are other research groups that will use the instrument to monitor processes such as RNA pseudoknot folding, ligand binding-induced changes in circular dichroism and/or formation of transient intermediates during enzyme catalyzed reactions. To date, virtually all published experiments monitoring CD on a stopped flow time scale have been to study protein folding, and it is clear that much of the time on the proposed instrument will be devoted to this application. However, based on our experience with CD to monitor binding reactions at equilibrium, it is clear that access to the instrumentation will allow us to monitor kinetics of processes other than protein folding. Specifically, chromophores such as flavins, pterins and some metal complexes exhibit large changes in circular dichroisn in response to changes in local environment that may be diagnostic of mechanistically important processes. Thus, while the instrument can be fully justified based on our current activities in protein folding, we fully anticipate developing additional applications that will add to its value within the user community. The faculty participants in this project have a long history of productive interaction and collaborations. Through the Center for Macromolecular Design, which is sponsoring this proposal, we have in place a series of shared instrumentation facilities, including a Varian Unity Plus 500 MHz NMR Spectrometer, a Beckman Optima XLA analytical ultracentrifuge, a Laboratory for Biological Mass Spectrometry, and a Protein Chemistry Laboratory. These multidepartmental, multicollege facilities are strongly supported by the administration of the university, and provide a cost- effective mechanism to provide state-of-the-art instrumentation and expertise for research programs on the campus of Texas A&M University.
