Protein dynamics of calcium-S100A5 in the presence and absence of target peptide

存在和不存在目标肽时钙-S100A5 的蛋白质动力学

• 批准号: 7983430
• 负责人: Melissa A. Liriano
• 金额: $ 3.87万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-11-16 至 2012-11-15
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7983430
• 关键词: Affinity; Amides; Astrocytoma; Binding; Biological; Biological Process; Calcium; Calcium Binding; Calcium Channel; Cardiomyopathies; Characteristics; Chemicals; Complex; Comprehension; Data; Disease; Drug Design; EF Hand Motifs; Family; Family member; Goals; Homeostasis; Individual; Inflammation; Ions; Label; Ligands; Link; Malignant Neoplasms; Measurement; Melissa; Metabolism; Molecular Structure; Molecular Target; Neurons; Normal Cell; Parvalbumins; Peptides; Physiological; Physiological Processes; Play; Positioning Attribute; Process; Protein Dynamics; Protein Family; Proteins; Relative (related person); Relaxation; Role; S100 Proteins; Side; Signaling Molecule; Sodium; Source; Structure; Testing; Time; Vertebral column; Vertebrates; X-Ray Crystallography; base; cell growth; cell type; drug development; gene therapy; insight; interest; melanoma; member; meningioma; methyl group; nervous system disorder; novel; novel therapeutics; peptide structure; programs; protein protein interaction; research study; theories; tumor; tumor progression

DESCRIPTION (provided by applicant): The SI 00 family of proteins are involved in many physiological processes. Aberrant levels ofthese signaling molecules have also been associated with various forms of cancer. Some S100 proteins have been found to directly promote the advancement of certain tumors via protein-protein interactions. The long- term objective ofthis proposal is to study the calcium-dependent interaction between S100 proteins and their molecular targets. A majority of SI 00 members bind calcium with a low binding affinity, unless a target is present. However, S100A5, an SI00 protein associated with recurring meningiomas and some astrocytomas, binds calcium with an affinity 10-100 times greater than any other SI 00 protein, independent of target. Determining the source of SI OOAS's high affinity for calcium in the absence of target can give us insight into how these molecules interact with targets in physiological and pathophysiological processes (ie. tumor progression). We have preliminary evidence to suggest that S100A5 lacks the protein dynamics common to the SI 00 family which increases its affinity for calcium. The goal of Aim 1 is to solve the structure of calcium-Si 00A5 with X-ray crystallography. Of particular interest is the calcium coordination in SI OOAS's canonical EF-hand which may be unlike most SI 00 proteins. An extra ligand at the ninth position ofthe calcium coordinating sphere, much like the protein parvalbumin, may potentially enhance calcium binding and we intend to determine this directly with X-ray crystallography. In aim 2, we will solve the structure of S100A5 bound to a peptide derived from the intracellular regulatory loop ofthe sodium-calcium channel (NCX) using NMR. An SIOOAS-target complex structure can give important information regarding general dynamic characteristics ofthe complex from multiple NMR parameters, such as NOEs or chemical exchange (ex. Rex). The SIOOAS-peptide structure also has biological significance in neurons. We predict that S100A5 plays a role in calcium homeostasis by directly competing with CaM to bind NCX thus inhibiting calcium efflux. Our final aim is to compare the protein dynamics of S100A5 +/- target to known low (ex. SIOOB-calcium) and high (ex. SIOOB-calcium+target) calcium affinity states. Through the use of specific 15N and 2H NMR relaxation measurements, we can directly ascertain the dynamics of backbone and side chain amide groups and methyl-bearing side chains on a variety of time scales, from ms to ps. Since SI 00 proteins are elevated in multiple disease states, comprehension ofthe molecular interactions between SI 00 proteins and targets can provide objectives for novel therapeutics, such as the ongoing drug design program in the Weber lab for inhibiting S100B as a means to treat malignant melanoma.

描述（由申请人提供）：SI00蛋白质家族参与了许多生理过程。这些信号分子的异常水平也与各种形式的癌症有关。已经发现一些S100蛋白可以通过蛋白质蛋白质相互作用直接促进某些肿瘤的发展。这一建议的长期目标是研究S100蛋白质及其分子靶标之间的钙依赖性相互作用。除非存在靶标，否则大多数SI 00成员的结合亲和力与低结合亲和力结合。然而，S100A5是一种与反复出现的脑膜瘤和某些星形胶质细胞瘤相关的SI00蛋白，其亲和力比任何其他SI 00蛋白都高10-100倍，而与靶标无关。在没有目标的情况下，确定Si ooas对钙的高亲和力的来源可以使我们深入了解这些分子如何与生理和病理生理过程中的靶标相互作用（即肿瘤进展）。我们有初步证据表明，S100A5缺乏SI00家族共有的蛋白质动力学，从而增加了其对钙的亲和力。 AIM 1的目的是用X射线晶体学解决钙-SI 00A5的结构。特别令人感兴趣的是Si oa的规范EF手中的钙配位，这可能与大多数Si 00蛋白不同。像蛋白质白蛋白一样，在钙配位球体第九位的额外配体可能会增强钙结合，我们打算用X射线晶体学直接确定这一点。在AIM 2中，我们将使用NMR求解源自钠钙通道（NCX）的细胞内调节环的S100A5的结构。 SiOoAS-Target复合体可以从多个NMR参数（例如NOES或Chemical Exchange）（例如REX）提供有关复合物的一般动态特征的重要信息。 SiOoAS肽结构在神经元中也具有生物学意义。我们预测S100A5通过直接与CAM竞争结合NCX从而抑制钙外排在钙稳态中起作用。我们的最终目的是将S100A5 +/-靶标的蛋白质动力学与已知的低（sioob-calcium）和高（Ex。SiooOb-Calcium +Target）钙亲和力状态进行比较。通过使用特定的15N和2H NMR弛豫测量值，我们可以直接确定从MS到PS的各种时间尺度上的主链和侧链酰胺基和带甲基的侧链的动力学。由于在多种疾病状态下SI00蛋白升高，因此对SI00蛋白质和靶标之间的分子相互作用的理解可以为新型治疗剂提供目标，例如Weber Lab中正在进行的药物设计计划抑制S100B作为治疗恶性黑色素瘤的手段。