(1) Structural mechanism of multi-substrate selectivity in urea/amide channel (UAC) from Bacillus cereus UAC from B. cereus is highly permeable to both urea and nicotinamide, whose structures and sizes are different. This makes UAC an interesting target to study the mechanism of selectivity on multiple substrates. UAC is a hexameric membrane protein of 132 kDa with each protomer as an independent channel. It is fully embedded in the lipid membrane without a soluble domain. Its small size makes it a challenging target for cryoEM structural study. We have expressed and purified UAC from E. coli for structural and functional studies. Using single particle cryoEM, we have obtained the structures of UAC in the apo form and in complex with its substrates at atomic resolution. The structures suggest that UAC does not possess a fully open permeation pathway and instead it works like a transporter in which two key residues open and close alternatively to provide a mechanism of alternating access. (2) Structural study of anion exchanger 1 (AE1) AE1 (a.k.a. Band 3) is a member of the solute carrier (SLC) 4 family of bicarbonate transporters. It is the major membrane protein in red blood cells. It mediates fast exchange of bicarbonate and chloride ions across the cell membrane by its membrane domain and also participates in maintenance of the erythrocyte shape by binding to cytoskeletal proteins through its cytoplasmic domain. Mutations in AE1 are implicated in red cell diseases, including thalassemia, sickle cell anemia, and Southeast Asian ovalocytosis. We have used cryoEM to solve the structures of human AE1 membrane domain (about 120 kDa), which is the transporter domain, in both the inward-facing and the outward-facing conformations. Comparison between these two different opening states provides some interesting insights into the mechanism of fast transport conducted by AE1.

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National Heart, Lung, and Blood Institute
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