Characterization of water channel proteins (aquaporins) in plants.

摘要

Aquaporins are intrinsic membrane proteins which act as pores that are highly selective for water, or in some cases water and small solutes. These proteins therefore facilitate the osmotically- or hydraulically-driven bulk flow of water across certain biological membranes. Aquaporins belong to a group of proteins known as the MIP family, which has members in a diverse range of organisms ranging from bacteria to insects, plants and mammals. In plants, water channel proteins are thought to play a role in the systemic flow of water driven by transpiration, and to function in a number of important physiological processes, including cell expansion and tissue growth, maintenance of cell turgor, and seed rehydration. However, plant aquaporins are not well characterized and their involvement in these activities has yet to be established.; The goal of my dissertation research has been to characterize water channel proteins in plants. I have cloned and characterized the novel aquaporin {dollar}delta{dollar}-TIP from the tonoplast of Arabidopsis thaliana and characterized the Arabidopsis plasma membrane aquaporin RD28. Through mutational analysis I have discovered amino acid residues of {dollar}delta{dollar}-TIP, RD28, and the Arabidopsis tonoplast aquaporin {dollar}gamma{dollar}-TIP which are important in the pharmacological inhibition of water channel activity and provide insight into aquaporin structure. Transformed plants were generated which express RD28 and mutant RD28 aquaporins. Analysis of these plants may link aquaporins to water flow through plant tissues.; To determine the mechanism for channel substrate specificity in aquaporins and other MIP family proteins, I have purified, crystallized, and begun the structural characterization of the Phaseolus vulgaris aquaporin {dollar}alpha{dollar}-TIP. By electron and cryo-electron microscopy of {dollar}alpha{dollar}-TIP crystals I am working towards solving the atomic structure of this plant aquaporin. Crystals of {dollar}alpha{dollar}-TIP and the data derived from them show that this aquaporin will share features with previously crystallized aquaporins but may allow the resolution of water channel structure to a degree not previously obtainable.