INTRINSICALLY DISORDERED PROTEINS IN BIOMINERALIZATION

生物矿化中的本质无序蛋白质

• 批准号: 8119445
• 负责人: Janet M. Oldak
• 金额: $ 38.62万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-25 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8119445
• 关键词: 3-Dimensional; Affect; Amelogenesis Imperfecta; Amino Acid Sequence; Amino Acids; Apatites; Binding; Biomimetic Materials; C-terminal; Calcium; Calcium Carbonate; Cell Communication; Cell Surface Proteins; Cell surface; Characteristics; Chemicals; Circular Dichroism; Classification; DSPP gene; Dental; Dental Enamel; Dentin; Dentin Formation; Development; Disease; Enamel Formation; Environment; Equilibrium; Event; Excision; Extracellular Matrix; Extracellular Matrix Proteins; Extracellular Protein; Family suidae; Fluorescence; Genes; Goals; Grant; Growth; Investigation; Minerals; Molecular; Molecular Conformation; Monitor; Mutation; NMR Spectroscopy; Natural regeneration; Nature; Oral; Outcome; Peptide Hydrolases; Peptide Sequence Determination; Play; Prevalence; Process; Property; Proteins; Proteolysis; Recombinants; Regulation; Reporting; Research; Role; Series; Shapes; Signal Transduction; Solutions; Spectroscopy, Fourier Transform Infrared; Structure; Surface; System; Techniques; Testing; Tissues; Titrations; Tooth structure; amelogenin; biomineralization; bone; calcium phosphate; conformational conversion; craniofacial; enamelin; flexibility; improved; knockout animal; mineralization; novel; polypeptide; programs; protein folding; proteinase In; public health relevance; repaired; self assembly; synthetic peptide; three dimensional structure; tissue regeneration

DESCRIPTION (provided by applicant): This proposal will explore novel structural principles by which proteins interact with their targets and will investigates the prevalence of "intrinsically disordered" proteins (IDPs) in the field of tooth biomineralization. The long term objective of the proposed research is to advance understanding of the role of extracellular matrix protein fragmentation in enamel and dentin, with a focus on the analysis of folding and the degree of disorder in the secondary structures of key proteins and polypeptides. We propose that disordered domains in the extracellular matrix proteins play substantial roles in biomineralization. We hypothesize that the programmed proteolytic activities in enamel and dentin regulate protein-mineral, protein-protein and protein- cell interactions through regulation of the number of unstructured regions and the degree of disorder in the protein sequence. Such interactions will affect chemical and cellular events such as cell signaling, macromolecular self-assembly, crystal nucleation and growth, and protein removal. Three specific aims are proposed: Aim I. To apply computational and biophysical strategies to analyze the secondary structures and evaluate the degree and nature of "disorder" in key proteins of the extracellular matrix of enamel and dentin as well as their selected proteolytic fragments. Aim II: To use circular dichroism (CD) and fluorescence titrations to determine the conformational changes and strength of interactions between key fragments of amelogenin and ameloblastin, as well as DPP with their macromolecular targets. To use NMR spectroscopy to monitor key amino acid residues that undergoes conformational transitions in amelogenin sequence as the result of binding to partners/targets. Aim III: To use spectroscopical techniques (CD and ATR-FTIR) to monitor secondary structural changes and conformational transitions of selected amelogenin, enamelin, and ameloblastin proteolytic fragments, as well as DPP following their interactions with calcium and with apatite crystal surfaces. In Summary: Enamel and dentin biomineralization is the result of orchestration among a series of protein-protein, protein-mineral and protein cell interactions. Our goal is to systematically dissect the sequence and secondary structures of functional domains in major enamel extracellular matrix proteins as well as the C-terminal portion of dentin sialophosphoprotein (DPP) with regard to IDPs, and to provide information on secondary structural alteration as the result of protein-protein, protein-mineral and protein-cell surface interactions. Understanding the role of "intrinsic disorder" in proteins of extracellular matrix of enamel and dentin will prepare the ground for the fabrication and development of biomimetic materials when synthetic peptides can be used to control the processes of crystal nucleation and growth. Identification of unfolded functional domains in cell signaling will have a great impact in the field of tissue regeneration. The outcomes of our study will therefore have the potential to improve treatments for repair and regeneration of oral, dental and craniofacial tissues. PUBLIC HEALTH RELEVANCE: This proposal will explore novel structural principles by which proteins interact with their targets and will investigates the prevalence of "intrinsically disordered" proteins (IDPs) in the field of tooth biomineralization. The outcomes will prepare the ground for the fabrication and development of biomimetic materials when synthetic peptides can be used to control the processes of crystal nucleation and growth. Identification of unfolded functional domains in cell signaling will have a great impact in the field of tissue regeneration. The outcomes of our study will therefore have the potential to improve treatments for repair and regeneration of oral, dental and craniofacial tissues.

描述（由申请人提供）：该提案将探索蛋白质与目标相互作用的新型结构原理，并将研究牙齿生物矿化领域中“本质上无序”蛋白（IDP）的患病率。拟议的研究的长期目标是提高对牙釉质和牙本质中细胞外基质蛋白碎片作用的了解，重点是分析折叠和关键蛋白质和多肽的二级结构中的疾病程度。我们提出，细胞外基质蛋白中的无序结构域在生物矿化中起着重要作用。我们假设在牙釉质和Dentin中的编程蛋白水解活性通过调节非结构化区域的数量以及蛋白质序列中的疾病程度来调节蛋白质矿物质，蛋白质蛋白质和蛋白质细胞相互作用。这种相互作用将影响化学和细胞事件，例如细胞信号传导，大分子自组装，晶体成核和生长以及去除蛋白质。提出了三个具体目的：目标I.应用计算和生物物理策略来分析二级结构并评估搪瓷和牙本质细胞外基质的关键蛋白质中的“疾病”的程度和性质，以及其选定的蛋白水解片段。 AIM II：使用圆形二色性（CD）和荧光滴定来确定氨基蛋白蛋白和氨基蛋白细胞蛋白和杏仁糖蛋白关键片段之间相互作用的构象变化和强度，以及与大分子靶标的DPP。使用NMR光谱法监测关键氨基酸残基，该氨基酸残基在与伴侣/靶标结合的结果中在氨基蛋白蛋白序列中进行构象转变。 AIM III：使用光谱技术（CD和ATR-FTIR）来监测所选氨基蛋白蛋白，搪瓷蛋白和蛋白蛋白蛋白水解蛋白水解片段的二级结构变化和构象转变，以及与钙与钙层晶体相互作用后的DPP以及DPP。总结：搪瓷和牙本质生物矿化是一系列蛋白质 - 蛋白质，蛋白质矿物质和蛋白质细胞相互作用之间编排的结果。我们的目标是系统地剖析主要牙釉质外基质基质蛋白中功能域的序列和二级结构，以及dentin sialophophopolotein（DPP）的C末端部分，与IDP有关，并提供有关二次结构变化的信息，以提供蛋白质 - 蛋白质 - 蛋白质 - 蛋白质中的蛋白质，蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 蛋白质 - 中等。当可以使用合成肽可用于控制晶体成核和生长过程时，了解搪瓷和牙本质细胞外基质的蛋白质中的作用将为仿生材料的制造和发育做好准备。细胞信号传导中未折叠的功能结构域的鉴定将在组织再生领域产生重大影响。因此，我们研究的结果将有可能改善口服，牙齿和颅面组织修复和再生的治疗方法。 公共卫生相关性：该建议将探讨蛋白质与目标相互作用的新型结构原理，并将研究牙齿生物矿化领域中“本质上无序”蛋白（IDP）的流行。当合成肽可用于控制晶体成核和生长的过程时，结果将为制造和开发仿生材料的制造和开发做准备。细胞信号传导中未折叠的功能结构域的鉴定将在组织再生领域产生重大影响。因此，我们研究的结果将有可能改善口服，牙齿和颅面组织修复和再生的治疗方法。