Browsing by Subject "oocyte electrophysiology"

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    Structure-function relationship of plant sucrose transporters (SUTs)
    (2012-07) Sun, Ye
    Sucrose transporters (SUTs or SUCs) are membrane proteins that transport sucrose and H+ into the cytoplam at a ratio of 1:1. They are important for the long-distance transport of sucrose in plants. However, little is known about the structure-function relationship of SUTs. In this thesis, the transport activity and substrate specificity of rice SUTs were measured using [14C] sucrose yeast uptake and oocyte electrophysiology. More importantly, a 3D structural model of rice sucrose transporter OsSUT1 was built using known crystal structures of transporters from E.coli as templates. Based on the predicted model, six charged amino acids in transmembrane spans were selected for mutagenesis, five of which turned out to be essential for the SUT transport function. One mutant, R188K, caused a complete loss of sucrose transport activity, and showed a H+ leak that could be blocked by sucrose. Based on electrophysiology experiments results, a putative binding interaction between Arg188 of OsSUT1 and hydroxyl groups of sucrose was proposed. A role of Arg188 in the substrate transport process was also suggested. In addition, methods to identify amino acids important for SUT substrate specificity were explored.