Previous studies have established that the folding, structure and function of membrane proteins are influenced by their lipid environments1-7 and that lipids can bind to specific sites, for example in potassium channels8. Fundamental questions remain however regarding the extent of membrane protein selectivity toward lipids. Here we report a mass spectrometry (MS) approach designed to determine the selectivity of lipid binding to membrane protein complexes. We investigate the mechanosensitive channel of large conductance (MscL), aquaporin Z (AqpZ), and the ammonia channel (AmtB) using ion mobility MS (IM-MS), which reports gas-phase collision cross sections. We demonstrate that folded conformations of membrane protein complexes can exist in the gas-phase. By resolving lipid-bound states we then rank bound lipids based on their ability to resist gas phase unfolding and thereby stabilize membrane protein structure. Results show that lipids bind non-selectively and with high avidity to MscL, all imparting comparable stability, the highest-ranking lipid however is phosphatidylinositol phosphate, in line with its proposed functional role in mechanosensation9. AqpZ is also stabilized by many lipids with cardiolipin imparting the most significant resistance to unfolding. Subsequently, through functional assays, we discover that cardiolipin modulates AqpZ function. Analogous experiments identify AmtB as being highly selective for phosphatidylglycerol prompting us to obtain an X-ray structure in this lipid membrane-like environment. The 2.3Å resolution structure, when compared with others obtained without lipid bound, reveals distinct conformational changes that reposition AmtB residues to interact with the lipid bilayer. Overall our results demonstrate that resistance to unfolding correlates with specific lipid-binding events enabling distinction of lipids that merely bind from those that modulate membrane protein structure and/or function. We anticipate that these findings will be influential not only for defining the selectivity of membrane proteins toward lipids but also for understanding the role of lipids in modulating function or drug binding.

Amyloid diseases, including Alzheimer's, Parkinson's, and the prion conditions, are each associated with a particular protein in fibrillar form. These amyloid fibrils were long suspected to be the disease agents, but evidence suggests that smaller, often transient and polymorphic oligomers are the toxic entities. Here we identify a segment of the amyloid-forming protein, alphaB crystallin, which forms an oligomeric complex exhibiting properties of other amyloid oligomers: beta-sheet-rich structure, cytotoxicity, and recognition by an anti-oligomer antibody. The X-ray-derived atomic structure of the oligomer reveals a cylindrical barrel, formed from six anti-parallel, protein strands, which we term a cylindrin. The cylindrin structure is compatible with a sequence segment from the Abeta protein of Alzheimer's disease. Cylindrins offer models for the hitherto elusive structures of amyloid oligomers.

Amyloid-beta (Aβ) aggregates are the main constituent of senile plaques, the histological hallmark of Alzheimer's disease. Aβ molecules form β-sheet containing structures that assemble into a variety of polymorphic oligomers, protofibers, and fibers that exhibit a range of lifetimes and cellular toxicities. This polymorphic nature of Aβ has frustrated its biophysical characterization, its structural determination, and our understanding of its pathological mechanism. To elucidate Aβ polymorphism in atomic detail, we determined eight new microcrystal structures of fiber-forming segments of Aβ. These structures, all of short, self-complementing pairs of β-sheets termed steric zippers, reveal a variety of modes of self-association of Aβ. Combining these atomic structures with previous NMR studies allows us to propose several fiber models, offering molecular models for some of the repertoire of polydisperse structures accessible to Aβ. These structures and molecular models contribute fundamental information for understanding Aβ polymorphic nature and pathogenesis.amyloid aggregation | 3D profile | protofilaments | heterotypic zipper