Protein interactions by analytical ultracentrifugation

描述（由申请人提供）：芝加哥伊利诺伊大学结构生物学中心（UIC）提议收购Beckman Coulter XL-I分析超离心机及其附件，包括Aviv Biomedical新提供的荧光检测系统，以推进美国国立卫生研究院资助的五个研究小组的研究，所有这些研究都涉及蛋白质与其他蛋白质或配体的相互作用研究。这些设备将被安置在结构生物学中心的共用设备室内。主要用户为我校医学、药学、牙科三个学院的生物化学系、药物生物技术中心、口腔疾病分子生物学中心。该设备将补充目前可用的共享仪器，包括超高场核磁共振、荧光、CD光谱仪、DSC和ITC量热仪，通过对感兴趣的蛋白质与其他蛋白质和配体的溶液相互作用进行定量检查，而不管相互作用中是否存在光谱变化或可测量的焓变化。荧光检测能力将显著扩展xl - 1在可检测系统范围方面的能力，包括可测量的解离常数范围、可分析系统的复杂性和样品节约。将研究的系统包括(i) HIV病毒包膜蛋白gp41和gp120以及SARS冠状病毒的S1和S2 (Caffrey), (ii) LDL受体相关蛋白与其伴侣RAP和蛇形蛋白蛋白酶配体的相互作用，以及人类泛蛋白酶抑制剂α -2-巨球蛋白（Gettins）结构域-结构域相互作用的表征，（iii）药物前加工酶调控中的蛋白-蛋白相互作用。以及支架蛋白在特定细胞位点组织多个伙伴的作用（Lavie）， （iv）抗氧化反应中的异质相互作用，3类去乙酰化酶、炭疽芽孢杆菌代谢途径酶和SARS冠状病毒蛋白酶的寡聚化状态，这些酶将被新型蛋白酶抑制剂（Mesecar）的开发所抑制，以及(v)抗血管生成形式的抗凝血酶的构象变化。crmA-caspase复合物的组织，以及ZPI、Z蛋白和Xa因子之间形成的复合物的研究，该复合物允许ZPI抑制Xa因子（Olson）。使用这种设备进行的研究将解决与人类病毒感染（如HIV或SARS）或血液凝固等正常生理过程有关的蛋白质的结构和功能的基本问题。所获得的知识对于设计针对此类感染的治疗方法或改善与血液凝固等过程异常功能相关的疾病将是无价的。

DESCRIPTION (provided by applicant): The Center for Structural Biology at the University of Illinois at Chicago (UIC) proposes acquisition of a Beckman Coulter XL-I analytical ultracentrifuge, with accessories, including the newly-available Fluorescence Detection System from Aviv Biomedical, to advance the research of five NIH-funded research groups, all of which involve the study of protein interactions with other proteins or ligands.The equipment will be housed in the Center for Structural Biology in a shared equipment room. The primary users belong to three departments (Biochemistry, Center for Pharmaceutical Biotechnology, and the Center for Molecular Biology of Oral Disease) in three colleges (Medicine, Pharmacy and Dentistry) of the university. The equipment will complement currently available shared instrumentation that includes ultra high field NMR, fluorescence, and CD spectrometers and DSC and ITC calorimeters by enabling quantitative examination of the solution interactions of proteins of interest with other proteins and ligands, irrespective of whether there are spectroscopic changes or measurable enthalpy changes in the interactions. Fluorescence detection capabilities will significantly extent the capabilities of the XL-I in terms of the range of systems that can be examined, including range of dissociation constants that can be measured, the complexity of systems analyzable, and sample sparing. Systems that will be studied include (i) viral envelope proteins gp41 and gp120 from HIV and the S1 and S2 from SARS coronavirus (Caffrey), (ii) interactions of the LDL receptor related protein with its chaperone RAP and with serpin-proteinase ligands, and characterization of domain-domain interactions in the human pan-proteinase inhibitor alpha-2-macroglobulin (Gettins), (iii) protein-protein interactions in the regulation of pro-drug processing enzymes, and in the role of scaffold proteins in organizing multiple partners at specific cellular sites (Lavie), (iv) heterogeneous interactions in the antioxidant response, the oligomerization state of class 3 deacetylases, of Bacillus anthracis metabolic pathway enzymes, and of proteases from the SARS coronavirus that will be targeted for inhibition by development of novel protease inhibitors (Mesecar) and (v) conformational changes in antithrombin in the antiangiogenic forms, organization of crmA-caspase complexes, and studies on the complex formed between ZPI, protein Z and factor Xa that permits inhibition of factor Xa by ZPI (Olson). The research to be carried out using this equipment addresses basic questions of the structure and function of proteins that are involved in human viral infections such as HIV or SARS, or normal physiological processes such as blood coagulation. The knowledge gained will be invaluable for design of therapeutics against such infections or in amelioration of diseases associated with aberrant functioning of processes such as blood coagulation.