Department of Food Science and Technology, South China University of Technology, Guangzhou 510640, People's Republic of China.
J Agric Food Chem. 2010 Jul 14;58(13):8061-8. doi: 10.1021/jf101311f.
The amyloid fibrils formed by heating 1.0% (w/v) kidney bean phaseolin (7S globulin) solution at pH 2.0 with an ionic strength of 20 mM at 85 degrees C were characterized using transmission electron microscopy (TEM), atomic force microscopy (AFM), binding of thioflavin T (Th T) and Congo Red dyes, and circular dichroism spectroscopy. The morphology of the formed fibrils was closely dependent upon heating time from 15 to 720 min. The diameters of the fibrils formed at various times were similar, but the mean contour length progressively increased with heating time. The Th T maximum fluorescence also progressively increased with heating time. The heating process caused remarkable changes in secondary, tertiary, and quaternary conformations of the phaseolin, but the extents of the changes were closely related to the heating time. With a short heating time (e.g., 15 min), the beta-strand content decreased from 38.7 to 22.9%, but further heating resulted in recovery of beta-strand structure. The tertiary and quaternary conformations gradually became flexible and unfolded upon heating. Gel electrophoresis analysis indicated that heating disrupted the polypeptides of phaseolin, leading to the formation of fragments with lower molecular mass (e.g., <10 kDa after 360 min). The results suggest that the amyloid fibril formation of phaseolin (7S globulin) involved the disruption of its polypeptides, as well as conformational changes at secondary, tertiary, and quaternary structural levels. This appears to be the first direct observation of amyloid fibrils from legume 7S storage globulin.
在 pH 2.0、离子强度为 20 mM 的条件下，将 1.0%（w/v）菜豆球蛋白（7S 球蛋白）溶液加热至 85°C，形成的淀粉样纤维通过透射电子显微镜（TEM）、原子力显微镜（AFM）、硫代黄素 T（Th T）和刚果红染料结合以及圆二色性光谱进行了表征。形成的纤维的形态与从 15 分钟到 720 分钟的加热时间密切相关。在不同时间形成的纤维的直径相似，但平均轮廓长度随着加热时间的增加而逐渐增加。Th T 最大荧光也随着加热时间的增加而逐渐增加。加热过程导致菜豆球蛋白的二级、三级和四级构象发生显著变化，但变化的程度与加热时间密切相关。在短的加热时间（例如 15 分钟）下，β-折叠含量从 38.7%下降到 22.9%，但进一步加热导致β-折叠结构的恢复。三级和四级构象随着加热逐渐变得灵活和展开。凝胶电泳分析表明，加热破坏了菜豆球蛋白的多肽，导致形成具有较低分子量的片段（例如，360 分钟后 <10 kDa）。结果表明，菜豆球蛋白（7S 球蛋白）的淀粉样纤维形成涉及其多肽的破坏，以及二级、三级和四级结构水平的构象变化。这似乎是首次直接观察到来自豆科 7S 贮藏球蛋白的淀粉样纤维。
