Department of Biochemistry and Biophysics, Center for Biomembrane Research, The Arrhenius laboratory, Stockholm University, Stockholm, Sweden.
FEBS J. 2010 Sep;277(18):3759-68. doi: 10.1111/j.1742-4658.2010.07777.x. Epub 2010 Aug 3.
The formation and structure of proinsulin C-peptide oligomers has been investigated by PAGE, NMR spectroscopy and dynamic light scattering. The results obtained show that C-peptide forms oligomers of different sizes, and that their formation and size distribution is altered by salt and divalent metal ions, which indicates that the aggregation process is mediated by electrostatic interactions. It is further demonstrated that the size distribution of the C-peptide oligomers, in agreement with previous studies, is altered by insulin, which supports a physiologically relevant interaction between these two peptides. A small fraction of oligomers has previously been suggested to be in equilibrium with a dominant fraction of soluble monomers, and this pattern also is observed in the present study. The addition of modest amounts of sodium dodecyl sulphate at low pH increases the relative amount of oligomers, and this effect was used to investigate the details of both oligomer formation and structure by a combination of biophysical techniques. The structural properties of the SDS-induced oligomers, as obtained by thioflavin T fluorescence, CD spectroscopy and IR spectroscopy, demonstrate that soluble aggregates are predominantly in β-sheet conformation, and that the oligomerization process shows characteristic features of amyloid formation. The formation of large, insoluble, β-sheet amyloid-like structures will alter the equilibrium between monomeric C-peptide and oligomers. This leads to the conclusion that the oligomerization of C-peptide may be relevant also at low concentrations.
已通过 PAGE、NMR 光谱和动态光散射研究了胰岛素原 C 肽寡聚物的形成和结构。研究结果表明，C 肽形成不同大小的寡聚物，并且其形成和大小分布受盐和二价金属离子的影响，这表明聚集过程受静电相互作用介导。进一步表明，C 肽寡聚物的大小分布与先前的研究一致，被胰岛素改变，这支持这两种肽之间存在生理相关的相互作用。以前曾提出，寡聚物的一小部分与占主导地位的可溶性单体的一部分处于平衡状态，本研究也观察到了这种模式。在低 pH 值下添加适量的十二烷基硫酸钠会增加寡聚物的相对含量，并且该效应被用于通过组合使用生物物理技术来研究寡聚物形成和结构的细节。通过硫代黄素 T 荧光、CD 光谱和红外光谱获得的 SDS 诱导的寡聚物的结构特性表明，可溶性聚集体主要处于β-折叠构象，并且寡聚化过程显示出淀粉样形成的特征。大的、不溶性的β-折叠样淀粉样结构的形成会改变单体 C 肽和寡聚物之间的平衡。这得出的结论是，C 肽的寡聚化在低浓度下也可能相关。
