Structural Studies of Sugar Transporters

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Membrane transport, for example glucose uptake, Is essential to the cell. Such processes are often mediated by secondary membrane transporters. The largest secondary transporter family is the major facilitator superfamily (MFS), with more than 5,000 members identified to date, including the glucose transporter from muscle (Glut4) and the glucose-6-phosphate transporter (G6PT), both from human and both are involved in the pathogenesis of type 2 diabetes. We propose to study the transport mechanisms of a bacterial homolog of Glut4 and G6PT, the glycerol-3-phosphate transporter (GIpT) from E. coli, using structural biology, biochemistry and spectroscopic approaches. In the previous funding cycle, we determined the crystal structure of GIpT in a substrate-free form at 3.3 A resolution. The structure suggests a "rocker-switch" mechanism for substrate transport. In the next funding cycle, to better understand GIpT's substrate specificity and substrate-induced conformational change, we plan to determine the crystal structures of the transporter in two substrate-bound forms. We will also test the rocker-switch mechanism and examine the role that key amino acid residues play using mutagenesis, transport assays and spectroscopic techniques. The structural and mechanistic information gained from GIpT can be used directly to improve our understanding of Glut4 and G6PT.