title: A cell-free translation and proteoliposome reconstitution system for functional analysis of plant solute transporters.
abstract: We describe here a novel proteoliposome reconstitution system for functional analysis of plant membrane transporters that is based on a modified wheat germ cell-free translation system. We established optimized conditions for the reconstitution system with Arabidopsis thaliana phosphoenolpyruvate/phosphate translocator 1 (AtPPT1) as a model transporter. A high activity of AtPPT1 was achieved by synthesis of the protein in the presence of both a detergent such as Brij35 and liposomes. We also determined the substrate specificities of three putative rice PPT homologs with this system. The cell-free proteoliposome reconstitution system provides a valuable tool for functional analysis of transporter proteins.
authors.affiliation: Cell-Free Science and Technology Research Center, and The Venture Business Laboratory, Ehime University, Matsuyama, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Nozawa
authors.forename: Hideaki
authors.initials: H
authors.lastname: Nanamiya
authors.forename: Takuji
authors.initials: T
authors.lastname: Miyata
authors.forename: Nicole
authors.initials: N
authors.lastname: Linka
authors.forename: Yaeta
authors.initials: Y
authors.lastname: Endo
authors.forename: Andreas P M
authors.initials: AP
authors.lastname: Weber
authors.forename: Yuzuru
authors.initials: Y
authors.lastname: Tozawa
mesh_terms: D017360:Arabidopsis / Q000378:metabolism*; D001483:Base Sequence; D002474:Cell-Free System; D017931:DNA Primers; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014176:Protein Biosynthesis; D011510:Proteolipids
chemical_list: D017931:DNA Primers; D026901:Membrane Transport Proteins; D011510:Proteolipids; C020485:proteoliposomes