## 1 Advances and Challenges in Proteomics  
Proteomics involves analysis of all the proteins from an organelle or cellular system. The data can include quantified protein levels, interactions, modifications, and turnover over a period of time or space. The information should be very accurate and reproducible given the high throughput data that is increasingly collected at multiple sites for clinical studies. Proteins can be classified as soluble, transmembrane, or peripheral membrane. Quantitative analyses of those that are membrane associated are particularly troublesome due to low abundance, hydrophobicity, and poor solubility issues, and hence, membrane proteome can be severely under-represented in proteomic studies [1; 2]. This is a concern as membrane proteins are intricately involved in disease mechanisms and represent the majority of drug targets. Proteomics analysis is usually carried out using mass spectrometry (MS) due to its high sensitivity and throughput. Both top-down and bottom-up approaches are employed. Top-down proteomics involves direct elucidation of intact proteins which are fractionated by gel electrophoresis or liquid chromatography (LC), and then fragmented and identified [3]. This approach is particularly useful for identifying isoforms and post-translational modifications (PTMs) of soluble proteins, less so for intact membrane proteins which are harder to separate in gels and columns. The technical challenges include generating minimally biased fractionation, ionization, and fragmentation of intact membrane protein complexes or memteins [4] for MS analysis in the gas phase. Bottom-up proteomics improves protein solubilization through the use of a denaturing detergent followed by digestion with a protease such as trypsin. The resulting peptides are then fractionated and identified based on MS-based fragmentation patterns. This approach reduces the complexity of the original heterogeneous mixtures and bypasses the problems associated with analyzing intact proteins through ionization and MS characterization. However, the detergents, such as sodium dodecyl sulfate (SDS), that are commonly used to solubilize or unfold the proteins to enhance protease access are also problematic. In particular, they compromise protease function and produce MS peak clusters that obscure peptide signals. Moreover, the buffers and salts used in the biological mixtures lead to matrix effects that cause MS signal suppression or enhancement. Alternative proteomic-based protocols that use liposomes rather than detergents face challenges due to the heterogeneity and hydrophobic crevasses in vesicles which tend to trap nonspecific interactors. Hence, improved sample preparation methods are desirable, particularly for removal of detergents and salts that compromise data read-out and that would allow accurate and reproducible analysis of membrane proteins. Proteomics protocols typically dissociate multimers and complexes with ligands including lipids, signaling molecules, and metabolites, particularly when detergents are used. Nonetheless, such interactions provide vital biological information, and ideally should be maintained during preparation. By analogy, one can count all the people in a room, but not know which ones are talking to one another let alone what they are saying. That is, proteomics can measure protein levels but not necessarily give insight into what the proteins are doing unless complexes can be kept intact to measure structures and kinetics. Hence, the preparative approaches should be compatible with parallel assays to characterize ligand interactions and high-resolution structures, thus yielding not only the players but also their partners and mechanisms. Detergent-based protocols confront proteomics and structural biology experts with problems that need to be addressed. Amongst the developments are the use of native nanodiscs to separate intact memteins.