Technische Universität Dortmund, Fakultät Physik and DELTA, D-44221 Dortmund, Germany.
Phys Chem Chem Phys. 2010 May 7;12(17):4375-82. doi: 10.1039/b925134k. Epub 2010 Feb 9.
A planar poly(acrylic acid) (PAA) brush provides an unusual substrate for the unspecific immobilization of proteins on material surfaces. At neutral pH-values, proteins adsorb at a PAA brush when the ionic strength of the protein solution is low. In contrast, raising the ionic strength to a few 100 mM transforms a PAA brush into a rather protein-resistant surface coating. Moreover, a PAA brush represents a mild environment for adsorbed proteins which preserves their secondary structure and biological activity. In this study, we focus on the biocompatibility of a PAA brush with an insulin solution. Insulin can form amyloid fibrils, which may also be triggered by interfaces. Using neutron reflectometry and attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy, the effects of pD value, ionic strength, and glycerol concentration on the density profile and the secondary structure of adsorbed insulin molecules at a PAA brush have been studied. At pD 7, insulin adsorbs at a PAA brush despite its negative net charge. As has been found for other proteins in earlier studies, increasing the ionic strength of the insulin solution to 500 mM decreases the amount of adsorbed insulin drastically. In contrast, at pD 2, addition of salt to the insulin solution induces a thick insulin adsorbate at a PAA brush suggesting both protein-brush and protein-protein interactions, i.e., insulin adsorption and aggregation to be effective. However, in the presence of 2 M glycerol, insulin adsorption is largely suppressed. Furthermore, no major alterations of the secondary structure of adsorbed insulin can be detected by ATR-FTIR spectroscopy under all conditions studied. Hence, the performed experiments demonstrate that a PAA brush does not promote the formation of insulin amyloid structures, which represents a fundamentally new aspect of the biocompatibility of this material surface coating.
一种平面的聚丙烯酸 (PAA) 刷为蛋白质在材料表面的非特异性固定提供了一个不寻常的基质。在中性 pH 值下，当蛋白质溶液的离子强度较低时，蛋白质会在 PAA 刷上吸附。相比之下，将离子强度提高到几百毫摩尔会将 PAA 刷转变为一种相当抗蛋白质吸附的表面涂层。此外，PAA 刷为吸附的蛋白质提供了一个温和的环境，保持其二级结构和生物活性。在这项研究中，我们专注于 PAA 刷与胰岛素溶液的生物相容性。胰岛素可以形成淀粉样纤维，这也可能被界面触发。使用中子反射谱和衰减全反射-傅里叶变换红外光谱 (ATR-FTIR)，研究了 pD 值、离子强度和甘油浓度对吸附在 PAA 刷上的胰岛素分子密度分布和二级结构的影响。在 pD 7 时，尽管胰岛素带有负净电荷，但仍会吸附在 PAA 刷上。正如在早期的研究中发现的其他蛋白质一样，将胰岛素溶液的离子强度增加到 500 mM 会大大减少吸附的胰岛素量。相比之下，在 pD 2 时，向胰岛素溶液中添加盐会导致 PAA 刷上形成厚的胰岛素吸附物，这表明存在蛋白质-刷和蛋白质-蛋白质相互作用，即胰岛素吸附和聚集是有效的。然而，在 2 M 甘油存在的情况下，胰岛素吸附被大大抑制。此外，在所有研究的条件下，ATR-FTIR 光谱都未检测到吸附的胰岛素二级结构的主要变化。因此，所进行的实验表明，PAA 刷不会促进胰岛素淀粉样结构的形成，这代表了这种材料表面涂层生物相容性的一个全新方面。
