9317845 O'Shea This proposal seeks funding for circular dicroism (CD) and UV VIS spectrophotometers. These instruments will be used in a series of biochemical and biophysical studies concerned with how proteins fold and how they interact with one another and with other macromolecules. The proposed research includes studies in the following broad research areas: (1) protein structure, structure prediction and protein folding, (2) protein protein and protein nucleic acid interactions. (1) Studies on protein structure, prediction and folding will involve both model peptides and a bacterial protease, a lytic protease. Computational predictions of protein structure will be tested using synthetic polypeptides. The structure of designed peptides will be studied to investigate fundamental aspects of protein stability and structure. Structural studies of a stable intermediate in the folding of a lytic protease will be carried out to understand the forces which stabilize this intermediate. Conversion of the intermediate to the native enzyme is catalyzed by an additional polypeptide sequence. Thus, a further goal of these studies is to understand the mechanism by which folding to the native state is catalyzed. (2) Studies on protein protein interactions will involve a group of proteins that play important roles in transcriptional regulation. These include the glucocorticoid receptor from mammalian cells and the SYMBOL 97 f "Symbol" 2, al, MCM1, PHO2, PHO4, and PHO80 proteins from yeast. A goal of these studies is to identify domains that comprise autonomous folding units, as well as domains that are responsible for inte ractions between proteins of multisubunit transcription complexes. An additional goal is to investigate changes in conformation of these molecules upon interaction with one another. These studies will provide an essential first step in obtaining three dimensional information on multi subunit protein complexes. The CD and UV VIS spectrophotometers are essential instruments for carrying out these studies. CD provides a simple and informative measure of secondary structure. It also provides a way to assess conformational changes in mixtures of macromolecules, such as two different proteins (or domains) or a protein and DNA. The UV VIS spectrophotometer provides a sensitive assay for measuring protein concentrations, a necessary component of a CD experiment. Additionally, UV VIS spectroscopy provides a method for measuring the stability of a molecule by monitoring absorbance. For proteins, the information derived from such a study is often complementary to that obtained from CD because changes in the UV spectrum are typically reflections of tertiary structure while CD is most sensitive to secondary structure. Finally, UV VIS spectroscopy provides a very sensitive way to monitor changes in nucleic acid stability and structure. These studies shall be carried out as part of ongoing, active research programs in which training of graduate students and postdoctoral fellows is a strong component. u ~ 9317845 O'Shea This proposal seeks funding for circular dicroism (CD) and UV VIS spectrophotometers. These instruments will be . / 0 x k m I K ' ) v x ! ! ! ! ! ! F x N x ; CG Times Symbol & Arial 1 Courier 0 MS LineDraw 9 N h = ABSTRACT Deseree King, BIR Deseree King, BIR
