PROTEIN BEHAVIOR AT SILICONE-WATER INTERFACES

硅-水界面上的蛋白质行为

• 批准号: 2186592
• 负责人: VIOLA VOGEL
• 金额: $ 9.76万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-01-01 至 1998-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2186592
• 关键词: acidity /alkalinity; acrylamides; adsorption; bilirubin; biomaterial interface interaction; chromophore; coagulation factor IX; complement; conformation; fibronectins; fluorescence microscopy; fluorescence spectrometry; intermolecular interaction; iodine; monoclonal antibody; oils; organosilicon compound; phosphatidylcholines; protein structure; serum albumin; thermodynamics; time resolved data; tryptophan; water

We propose to study conformational alternations of proteins adsorbed from solutions to water/silicone oil interfaces with the aim of assessing on the molecular level how silicone based implants, such as leaking breast implants, cause foreign body reactions. A long standing hypothesis is that the observed immunological responses do not originate from the silicone implants themselves, since silicones are characterized as chemically inert, but via proteins that undergo conformational alterations on surface adsorption. Protein adsorption to a synthetic surface occurs immediately after exposure; it hence constitutes the primary response. Little is known regarding protein conformation in the adsorbed state, not only on silicone oil surfaces but in general, since three-dimensional protein structures within adsorbate films can hardly be determined directly with monolayer sensitivity. It is therefore proposed to measure a set of physiochemical parameters for model proteins adsorbed to water/silicone interfaces, where each parameter probed reflects another feature of the conformationally altered protein, and then to compare these parameters for the same model protein adsorbed to other, far better characterized interfaces. It is assumed that if adsorbed proteins are similar in a sufficient number of physicochemical parameters they will express similar functional behavior. Experimentally, advantage is taken from the fact that silicone oils spread at the air/water interface. Protein behavior on adsorption to the water/silicone oil interface will hence directly be compared to that found at water/air and membrane mimetic water/lipid interfaces. This adsorption studies to all these interfaces will be done in a Langmuir trough. The uniqueness of this approach is that these three interfaces will be probed under similar conditions and by the same set of techniques which include measurements of surface potential, fluorescence spectroscopy and microscopy, time-resolved fluorescence anisotropy decay. The molecular ordering will be probed by a new powerful nonlinear optical laser technique, optical second harmonic generation. The model proteins are human serum albumin and human fibronectin. We will study their conformations at model interfaces and how they respond to solvent perturbation and pH (Experiment 1). The location of the tryptophan residues is deferred from probing their polar environment (Experiment 2), and their accessibility to aqueous quenches (Experiment 3). The conformational alterations will be related to the expressed function by studying the ability of surface adsorbed proteins to bind to ligands (Experiment 4). Information on the dynamics of the three- dimensional protein structure will be gained from time-resolved fluorescence anisotropy decay studies (Experiment 5). The texture of the adsorbed protein film will be imaged by fluorescence microscopy (Experiments 6), and the biological activity of the conformationally altered proteins will be assessed by studying their interaction with complement proteins and monoclonal antibodies (Experiment 7). The ultimate goal is to provide insight on the molecular level whether and how conformationally altered surface proteins may act indirectly as initiators of inflammatory responses.

我们建议研究蛋白质吸附的构象变化