Dissecting the Conformational Dynamics of the Bile Acid Transporter Homologue ASBT.

Apical sodium-dependent bile acid transporter (ASBT) catalyses uphill transport of bile acids using the electrochemical gradient of Na as the driving force. The crystal structures of two bacterial homologues ASBT and ASBT have previously been determined, with the former showing an inward-facing conformation, and the latter adopting an outward-facing conformation accomplished by the substitution of the critical Na-binding residue glutamate-254 with an alanine residue. While the two crystal structures suggested an elevator-like movement to afford alternating access to the substrate binding site, the mechanistic role of Na and substrate in the conformational isomerization remains unclear.

Site-Directed Alkylation Detected by In-Gel Fluorescence (SDAF) to Determine the Topology Map and Probe the Solvent Accessibility of Membrane Proteins.

The topology of helix-bundle membrane proteins provides low-resolution structural information with regard to the number and orientation of membrane-spanning helices, as well as the sidedness of intra/extra-cellular domains. In the past decades, several strategies have been developed to experimentally determine the topology of membrane proteins. However, generally, these methods are labour-intensive, time-consuming and difficult to implement for quantitative analysis.

Fluorophore Absorption Size Exclusion Chromatography (FA-SEC): An Alternative Method for High-Throughput Detergent Screening of Membrane Proteins.

Membrane proteins play key roles in many fundamental functions in cells including ATP synthesis, ion and molecule transporter, cell signalling and enzymatic reactions, accounting for ~30% genes of whole genomes. However, the hydrophobic nature of membrane proteins frequently hampers the progress of structure determination. Detergent screening is the critical step in obtaining stable detergent-solubilized membrane proteins and well-diffracting protein crystals.