Mass spectrometry of protein complexes - from networks to structures

点击翻译按钮获取中文摘要

Mass spectrometry is well established in the fields of proteomics and metabolomics. Much less established is its role in deciphering the overall architecture of protein complexes. Using a procedure whereby we begin with the intact assembly we have shown that we can decompose the assembly into its simplest building blocks, look at their overall shape and then integrate them into 3D models of the overall complex. In order to do this we first have to validate the method using complexes of known structure. We have collaborated with a number of groups worldwide to obtain complexes with a variety of different topological arrangements and structures. Through a systematic survey of these complexes we hope to determine whether or not this approach will be suitable for use in deciding the overall architecture of unknown protein complexes. A further stream of our research involves isolating complexes at endogenous expression levels using a modified isolation protocol in which a single subunit is tagged and complexes are purified using affinity methods. We propose to continue and consolidate very exciting preliminary data that we have obtained for complexes involved in translation and splicing of RNA. The first of this series of complexes consists of the elongation initiation factors. By comparing and contrasting both human and yeast complexes we hope to gain novel insight into their subunit interaction maps and overall architecture. Similarly for the complexes involved in splicing, both yeast and human complexes will be studied, either in collaboration with Olga and E Makarov or by isolation from yeast strains grown in-house.