Killian Manuela S, Taylor Adam J, Castner David G
Department of Materials Science and Engineering, Chair for Surface Science and Corrosion, Friedrich-Alexander-University of Erlangen-Nuremberg, Martensstr. 7, 91058 Erlangen, Germany.
National ESCA and Surface Analysis Center for Biomedical Problems (NESAC/BIO), Molecular Engineering and Sciences Institute, University of Washington, Seattle, Washington 98195 and Departments of Bioengineering and Chemical Engineering, University of Washington, Seattle, Washington 98195.
Biointerphases. 2018 Jun 29;13(6):06E401. doi: 10.1116/1.5031189.
Exposure of protein modified surfaces to air may be necessary in several applications. For example, air contact may be inevitable during the implantation of biomedical devices, for analysis of protein modified surfaces, or for sensor applications. Protein coatings are very sensitive to dehydration and can undergo significant and irreversible alterations of their conformations upon exposure to air. With the use of two compatible solutes from extremophilic bacteria, ectoine and hydroxyectoine, the authors were able to preserve the activity of dried protein monolayers for up to >24 h. The protective effect can be explained by the preferred exclusion model; i.e., the solutes trap a thin water layer around the protein, retaining an aqueous environment and preventing unfolding of the protein. Horseradish peroxidase (HRP) immobilized on compact TiO was used as a model system. Structural differences between the compatible solute stabilized and unstabilized protein films, and between different solutes, were analyzed by static time-of-flight secondary ion mass spectrometry (ToF-SIMS). The biological activity difference observed in a colorimetric activity assay was correlated to changes in protein conformation by application of principal component analysis to the static ToF-SIMS data. Additionally, rehydration of the denatured HRP was observed in ToF-SIMS with an exposure of denatured protein coatings to ectoine and hydroxyectoine solutions.
在一些应用中，蛋白质修饰表面暴露于空气中可能是必要的。例如，在生物医学设备植入过程中、蛋白质修饰表面分析或传感器应用中，与空气接触可能是不可避免的。蛋白质涂层对脱水非常敏感，暴露于空气中时其构象可能会发生显著且不可逆的改变。通过使用来自嗜极端微生物的两种相容性溶质——四氢嘧啶和羟基四氢嘧啶，作者能够将干燥蛋白质单层的活性保持长达24小时以上。这种保护作用可以用优先排阻模型来解释；即，溶质在蛋白质周围捕获一层薄水层，保持水环境并防止蛋白质展开。固定在致密TiO上的辣根过氧化物酶（HRP）被用作模型系统。通过静态飞行时间二次离子质谱（ToF-SIMS）分析了相容性溶质稳定和未稳定的蛋白质膜之间以及不同溶质之间的结构差异。通过对静态ToF-SIMS数据应用主成分分析，比色活性测定中观察到的生物活性差异与蛋白质构象变化相关。此外，在ToF-SIMS中观察到，将变性蛋白质涂层暴露于四氢嘧啶和羟基四氢嘧啶溶液中时，变性的HRP会发生复水。
