Sandras Florent, Pézolet Michel, Marion Didier, Grauby-Heywang Christine
Centre de Physique Moleculaire Optique et Hertzienne, 33405 Talence cedex, France.
Langmuir. 2009 Jul 21;25(14):8181-6. doi: 10.1021/la9005086.
The conformation of puroindoline-a (PIN-a), a protein extracted from wheat endosperm, in free-standing black films has been studied using confocal Raman spectroscopy. This protein is characterized by the presence in its sequence of a unique tryptophan (Trp)-rich domain and of five disulfide bridges stabilizing its three-dimensional structure. PIN-a is able to form free-standing films, which are very stable in time, because of its remarkable surface-active properties. These films become black in a few hours and are characterized by the presence of numerous aggregates. Their Raman spectra show major modifications of the PIN-a structure as compared to the solid form, such as the formation of beta-sheets or unordered structures, the modification of the environment of the Trp domain and, the conformation of disulfide bridges. These modifications are in agreement with an unfolding of the protein at the interfaces of the film and suggest that the Trp domain is involved in the aggregation. We have also studied the influence of increasing amounts of lysopalmitoylphosphatidylcholine (LPC) into the films. The direct observation of these mixed films shows that LPC inhibits the formation of PIN-a aggregates and that the conformation of PIN-a is strongly correlated to the LPC/PIN-a molar ratio. Raman spectroscopy also shows that PIN-a disturbs the highly organized arrangement of LPC molecules in the film.
利用共聚焦拉曼光谱研究了从小麦胚乳中提取的一种蛋白质——麦醇溶蛋白-a（PIN-a）在自支撑黑色薄膜中的构象。该蛋白质的特征在于其序列中存在一个独特的富含色氨酸（Trp）的结构域以及五个稳定其三维结构的二硫键。PIN-a能够形成自支撑薄膜，由于其显著的表面活性特性，这些薄膜在很长时间内都非常稳定。这些薄膜在几小时内会变黑，其特征是存在大量聚集体。与固态形式相比，它们的拉曼光谱显示PIN-a结构有重大变化，例如β-折叠或无序结构的形成、Trp结构域环境的改变以及二硫键的构象变化。这些变化与蛋白质在薄膜界面处的展开一致，表明Trp结构域参与了聚集过程。我们还研究了向薄膜中添加越来越多的溶血棕榈酰磷脂酰胆碱（LPC）的影响。对这些混合薄膜的直接观察表明，LPC抑制PIN-a聚集体的形成，并且PIN-a的构象与LPC/PIN-a摩尔比密切相关。拉曼光谱还表明，PIN-a扰乱了薄膜中LPC分子的高度有序排列。
