Structure, supramolecular association and interaction of patho-active presenilin and betaAPP domains by high resolution mass spectrometry

在这个项目中，详细的结构，相互作用和超分子协会的b-淀粉样蛋白前体蛋白（bAPP）和presinilin（s）（PS），阿尔茨海默病的两个关键分子，将使用蛋白质质谱的新方法（电喷雾/ ESI;基质辅助激光解吸/ MALDI）进行评估。特别是，傅里叶变换-ICR-MS将被用作一种主要的生物分析工具，为结构研究提供灵敏度和分辨率。将通过化学合成制备跨越Ab跨膜序列和PS跨膜跨越区6和7的bAPP多肽，以在假定的蛋白水解切割位点处包括序列突变。将MS与本实验室开发的质谱表位作图相结合，研究PS与Ab序列的相互作用。一种新的亲和蛋白质组-MS方法，使用微毛细管和芯片固定的抗体将被应用于识别细胞内的关联产品和组件的PS复合物。此外，合成的大Ab-含有多肽的PS和bAPP的病理生理相互作用的未来研究提供了合适的模型。

In this project the detailed structures, interactions and supramolecular association of b-amyloid precursor protein (bAPP) and presinilin(s) (PS), two key molecules for Alzheimer's disease, will be evaluated using new methods of protein mass spectrometry (electrospray / ESI; matrix-assisted laser desorption / MALDI). In particular, Fourier-transform-ICR-MS will be used as a major bioanalytical tool providing ultrahigh sensitivity and resolution for structural studies. bAPP polypeptides spanning the Ab transmembrane sequences, and PS transmembrane spanning regions 6 and 7 will be prepared by chemical synthesis to include sequence mutations at the presumed proteolytic cleavage site. Coupling of MS with the mass spectrometric epitope mapping developed in our laboratory will be employed to study interactions of PS with Ab sequences. A new affinity proteome-MS approach using micro-capillary- and chip-immobilised antibodies will be applied to identify intracellular association products and components of the PS complex. Furthermore, the synthesis of large Ab-containing polypeptides should provide suitable models for future studies of pathophysiological interactions of PS and bAPP.