ATOMIC FORCE MICROSCOPY OF BIOPOLYMERS

生物聚合物的原子力显微镜

• 批准号: 7955876
• 负责人: Catherine E. Costello
• 金额: $ 2.38万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7955876
• 关键词: Amino Acid Sequence; Amyloid; Amyloid Fibrils; Amyloid beta-Protein; Aspirate substance; Atomic Force Microscopy; Biology; Biophysics; Biopolymers; Boston; Calibration; Carbon Nanotubes; Collagen; Complement; Complex; Computer Retrieval of Information on Scientific Projects Database; Development; Devices; Doctor of Philosophy; Electronic Mail; Fatty acid glycerol esters; Funding; Goals; Grant; Image; Individual; Institution; Maintenance; Mass Spectrum Analysis; Measurement; Measures; Medicine; Molecular; Molecular Weight; Monitor; Mucins; North Carolina; Operative Surgical Procedures; Patients; Post-Translational Protein Processing; Property; Proteins; Proteoglycan; Recombinants; Research; Research Personnel; Resolution; Resources; Sampling; Source; Supervision; Telephone; Testing; Tube; United States National Institutes of Health; Universities; Variant; Weight; amyloid fibril formation; base; experience; glycosylation; improved; instrument; macromolecule; single molecule; synuclein; tool

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The BUSM MS Resource has undertaken atomic force microscopy to complement mass spectrometry studies of individual biopolymers of high molecular weight, including, e.g., proteoglycans, collagens and molecules having collagen-type domains, and other proteins that contain high levels of glycosylation, as well as non-covalent complexes. AFM is being used to estimate the overall weight and modular composition of such species, on the basis of extrapolations that should become possible after construction of calibration curves over the range accessible by mass spectrometry. The aim of this project is to use AFM to quickly estimate the MWs of large molecules and complexes as a supplemental tool for mass spectrometry in the high molecular weight range. This is achieved by using AFM to measure the molecular volume of single molecules of known MW as standards, extrapolating a curve using biopolymer standards at different MW, and determining the relation between molecular volume and MW. AFM is also being used to characterize amyloid fibrils, with the goal of relating fibril properties to variations in the amino acid sequence and to posttranslational modifications of the constituent proteins. By using purified patient samples as well as recombinant a-beta protein, the formation of amyloid fibrils under different conditions and protein/GAG interactions is being monitored in both this development project and in several collaborative projects (e.g., Nugent, Skinner, Spencer, Trinkaus-Randall). AFM images have also been recorded for amyloid fibrils obtained from fat aspirates of patients with primary amyloid disease.To improve resolution in AFM measurements, the use of carbon nanotubes is being explored and a device for mounting the tubes has been constructed and tested. Day-to-day operation, maintenance and supervision of the instrument are the responsibility of Dr. Hong, who completed his PhD in the Boston University Dept. of Cellular Biophysics with a focus on AFM studies of mucins. Dr. Ding, who has extensive experience in AFM of beta-amyloid and synuclein amyloid fibrils and various types of macromolecules, helped in the planning for this project and continues to provide advice and assistance to the BUSM investigators. Although he has relocated to North Carolina, he remains in email and phone contact.

该子项目是利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 BUSM MS资源已经进行了原子力显微镜，以补充高分子量的单个生物聚合物的质谱研究，包括，例如，蛋白聚糖、胶原和具有胶原型结构域的分子，以及含有高水平糖基化的其它蛋白质，以及非共价复合物。原子力显微镜正被用来估计这些物种的总重量和模块组成，外推的基础上，应该成为可能后，建设校准曲线的范围内可通过质谱。本项目的目的是利用原子力显微镜快速估计大分子和复合物的分子量，作为质谱在高分子量范围内的补充工具。这是通过使用AFM测量已知MW的单分子的分子体积作为标准，使用不同MW的生物聚合物标准外推曲线，并确定分子体积和MW之间的关系来实现的。原子力显微镜也被用来表征淀粉样蛋白原纤维，与原纤维的性质有关的氨基酸序列的变化和翻译后修饰的组成蛋白质的目标。 通过使用纯化的患者样本以及重组α-β蛋白，在本开发项目和几个合作项目中监测了不同条件下淀粉样纤维的形成以及蛋白质/GAG相互作用（例如，Nugent，Skinner，Spencer，Trinkaus-Randall）.从原发性淀粉样蛋白病患者的脂肪抽吸物中获得的淀粉样蛋白原纤维的AFM图像也被记录下来。为了提高AFM测量的分辨率，正在探索使用碳纳米管，并且已经构建和测试了用于安装管的装置。 仪器的日常操作、维护和监督由洪博士负责，他在波士顿大学医学系完成了博士学位。细胞生物物理学的重点是粘蛋白的AFM研究。 丁博士在β-淀粉样蛋白和突触核蛋白淀粉样蛋白纤维和各种类型的大分子的AFM方面拥有丰富的经验，他帮助规划了这个项目，并继续为BUSM研究人员提供建议和帮助。 虽然他已经搬到了北卡罗来纳州，但他仍然保持着电子邮件和电话联系。