Journal Article

A Cell-Free Translation and Proteoliposome Reconstitution System for Functional Analysis of Plant Solute Transporters

Akira Nozawa, Hideaki Nanamiya, Takuji Miyata, Nicole Linka, Yaeta Endo, Andreas P. M. Weber and Yuzuru Tozawa

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 48, issue 12, pages 1815-1820
Published in print December 2007 | ISSN: 0032-0781
Published online December 2007 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pcm150
A Cell-Free Translation and Proteoliposome Reconstitution System for Functional Analysis of Plant Solute Transporters

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  • Biochemistry
  • Molecular and Cell Biology
  • Plant Sciences and Forestry

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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.

Keywords: Cell-free translation system; Phosphoenolpyruvate; Proteoliposome; Transporter; Wheat germ

Journal Article.  3428 words.  Illustrated.

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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