title: Projection structure of a plant vacuole membrane aquaporin by electron cryo-crystallography.
abstract: The water channel protein alpha-TIP is a member of the major intrinsic protein (MIP) membrane channel family. This aquaporin is found abundantly in vacuolar membranes of cotyledons (seed storage organs) and is synthesized during seed maturation. The water channel activity of alpha-TIP can be regulated by phosphorylation, and the protein may function in seed desiccation, cytoplasmic osmoregulation, and/or seed rehydration. Alpha-TIP was purified from seed meal of the common bean (Phaseolus vulgaris) by membrane fractionation, solubilization in diheptanoylphosphocholine and anion-exchange chromatography. Upon detergent removal and reconstitution into lipid bilayers, alpha-TIP crystallized as helical tubes. Electron cryo-crystallography of flattened tubes demonstrated that the crystals exhibit plane group p2 symmetry and c222 pseudosymmetry. Since the 2D crystals with p2 symmetry are derived from helical tubes, we infer that the unit of crystallization on the helical lattice is a dimer of tetramers. A projection density map at a resolution of 7.7 A revealed that alpha-TIP assembles as a 60 A x 60 A square tetramer. Each subunit is formed by a heart-shaped ring comprised of density peaks which we interpret as alpha-helices. The similarity of this structure to mammalian plasma membrane MIP-family proteins suggests that the molecular design of functionally analogous and genetically homologous aquaporins is maintained between the plant and animal kingdoms.
authors.affiliation: Department of Cell Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
authors.forename: M J
authors.initials: MJ
authors.lastname: Daniels
authors.forename: M J
authors.initials: MJ
authors.lastname: Chrispeels
authors.forename: M
authors.initials: M
authors.lastname: Yeager
mesh_terms: D000595:Amino Acid Sequence; D020346:Aquaporins / Q000737:chemistry* / Q000302:isolation & purification / Q000648:ultrastructure*; D020285:Cryoelectron Microscopy; D003460:Crystallization; D019281:Dimerization; D007887:Fabaceae / Q000737:chemistry* / Q000166:cytology; D007106:Immune Sera; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000648:ultrastructure; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010940:Plant Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000648:ultrastructure; D010946:Plants, Medicinal; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D014617:Vacuoles / Q000737:chemistry*
chemical_list: D020346:Aquaporins; D007106:Immune Sera; D008565:Membrane Proteins; D010940:Plant Proteins; C078044:alpha-TIP protein, Phaseolus vulgaris