Acquisition of Circular Dichroism and UV-VIS Spectrophotometers

9317845 O'Shea 该提案为圆二色性 (CD) 和紫外可见分光光度计寻求资金。这些仪器将用于一系列生物化学和生物物理研究，涉及蛋白质如何折叠以及它们如何彼此相互作用以及与其他大分子相互作用。拟议的研究包括以下广泛的研究领域的研究：（1）蛋白质结构、结构预测和蛋白质折叠，（2）蛋白质蛋白质和蛋白质核酸相互作用。 (1) 蛋白质结构、预测和折叠的研究将涉及模型肽和细菌蛋白酶（一种裂解蛋白酶）。蛋白质结构的计算预测将使用合成多肽进行测试。将研究设计的肽的结构，以研究蛋白质稳定性和结构的基本方面。将进行裂解蛋白酶折叠中稳定中间体的结构研究，以了解稳定该中间体的力。中间体向天然酶的转化由额外的多肽序列催化。因此，这些研究的进一步目标是了解催化折叠至天然状态的机制。 (2)蛋白质蛋白质相互作用的研究将涉及一组在转录调控中发挥重要作用的蛋白质。这些包括来自哺乳动物细胞的糖皮质激素受体和来自酵母的SYMBOL 97\f“Symbol”2,a1、MCM1、PHO2、PHO4和PHO80蛋白。这些研究的目标是识别包含自主折叠单元的结构域，以及负责多亚基转录复合物蛋白质之间相互作用的结构域。另一个目标是研究这些分子在相互相互作用时构象的变化。这些研究将为获得多亚基蛋白质复合物的三维信息提供重要的第一步。 CD 和紫外可见分光光度计是进行这些研究的必备仪器。 CD 提供了一种简单且信息丰富的二级结构测量方法。它还提供了一种评估大分子混合物构象变化的方法，例如两种不同的蛋白质（或结构域）或蛋白质和 DNA。 UV VIS 分光光度计提供了一种用于测量蛋白质浓度的灵敏测定方法，这是 CD 实验的必要组成部分。此外，紫外可见光谱提供了一种通过监测吸光度来测量分子稳定性的方法。对于蛋白质来说，从此类研究中获得的信息通常与从 CD 中获得的信息是互补的，因为紫外光谱的变化通常是三级结构的反映，而 CD 对二级结构最敏感。最后，紫外可见分光光度计提供了一种非常灵敏的方法来监测核酸稳定性和结构的变化。 这些研究应作为正在进行的、活跃的研究项目的一部分进行，其中研究生和博士后研究员的培训是一个重要组成部分。 u ~ 9317845 O'Shea 该提案为圆二色性 (CD) 和紫外可见分光光度计寻求资金。这些仪器将是。 / 0 x k m I K') v x ! ！ ！ ！ ！ ！ F×N×； CG Times Symbol & Arial 1 Courier 0 MS LineDraw 9 N h = 摘要 Deseree King，BIR Deseree King，BIR

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