The ability to maintain interactions between soluble protein subunits in the gas phase of a mass spectrometer gives critical insight into the stoichiometry and interaction networks of protein complexes. Conversely, for membrane protein complexes in micelles, the transition into the gas phase usually leads to the disruption of interactions, particularly between cytoplasmic and membrane subunits, and a mass spectrum dominated by large aggregates of detergent molecules. We show that by applying nanoelectrospray to a micellar solution of a membrane protein complex, the heteromeric adenosine 5'-triphosphate (ATP)-binding cassette transporter BtuC2D2, we can maintain the complex intact in the gas phase of a mass spectrometer. Dissociation of either transmembrane (BtuC) or cytoplasmic (BtuD) subunits uncovers modifications to the transmembrane subunits and cooperative binding of ATP. By protecting a membrane protein complex within a n-dodecyl-beta-d-maltoside micelle, we demonstrated a powerful strategy that will enable the subunit stoichiometry and ligand-binding properties of membrane complexes to be determined directly, by precise determination of the masses of intact complexes and dissociated subunits.

Here we examined the gas-phase structures of two tetrameric membrane protein complexes by ion mobility mass spectrometry. The collision cross sections measured for the ion channel are in accord with a compact configuration of subunits, suggesting that the native-like structure can be preserved under the harsh activation conditions required to release it from the detergent micelle into the gas phase. We also found that the quaternary structure of the transporter, which has fewer transmembrane subunits than the ion channel, is less stable once stripped of detergents and bulk water. These results highlight the potential of ion mobility mass spectrometry for characterizing the overall topologies of membrane protein complexes and the structural changes associated with nucleotide, lipid, and drug binding.

pigments is of interest to diverse alternative solar energy technologies including photoelectrochemical cells, [4][5][6][7][8][9][10][11][12][13] biosensing, [14,15] photosensing, [16] molecular electronics, [7] and solar fuel synthesis. [17][18][19] Studies have focused in the main on Photosystem I from cyanobacteria [20][21][22] and the RC and RC-LH1 complexes from purple photosynthetic bacteria such as Rhodobacter (Rba.) sphaeroides [23][24][25][26] (Figure 1a,b). This latter organism is a popular source of photoproteins because it is possible to apply extensive protein engineering to its well-characterized RC, enabling high-yield expression and purification of proteins with specifically tailored properties or substantial modifications.A feature of natural photosystems is selective harvesting of certain regions of the solar spectrum, the most obvious illustration being the predominant green color of plant photosynthetic tissues that arises from relatively strong absorbance of red and blue light by chlorophyll and carotenoid pigments. As Rba. sphaeroides synthesizes bacteriochlorophyll (BChl) as its primary photosynthetic pigment its RC exhibits strong absorbance in the near-infrared between 700 and 950 nm, and in the near-UV below 420 nm, but its absorbance across the visible region is relatively weak (Figure 1c). A limitation in the use of this protein in device technologies is therefore suboptimal harvesting of light energy across much of the region where the solar radiation at the earth's surface is maximal, [27] and this limitation is manifest in action spectra of photocurrent density in photoelectrochemical cells based on Rba. sphaeroides pigment-proteins. [28][29][30][31][32][33] In this study we investigated directed self-assembly of conjugates between genetically engineered Rba. sphaeroides RCs and water-soluble cadmium telluride (CdTe) quantum dots (QDs). The tuneable optical properties of these semiconductor nanocrystals have been exploited in a variety of technologies including solar cells and diverse biological applications. [34][35][36] The particular QDs employed in the present work have broad absorbance across the visible spectrum and an emission band centered at 750 nm that overlaps with RC absorbance bands centered at 760 and 800 nm (Figure 1c). These QDs therefore were capable of acting as a synthetic light harvesting system for energy transfer [37,38] and charge separation [23][24][25][26] in the Rba. sphaeroides RC (Figure 1b).Photoreaction centers facilitate the solar energy transduction at the heart of photosynthesis and there is increasing interest in their incorporation into biohybrid devices for solar energy conversion, sensing, and other applications. In this work, the self-assembly of conjugates between engineered bacterial reaction centers (RCs) and quantum dots (QDs) that act as a synthetic light harvesting system is described. The interface between protein and QD is provided by a polyhistidine tag that confers a tight and specific binding and defines the geometry of the interactio...