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

Akira Nozawa | Zendy; Hideaki Nanamiya | Zendy; Takuji Miyata | Zendy; Nicole Linka | Zendy; Yaeta Endo | Zendy; Andreas P.M. Weber | Zendy; Yuzuru Tozawa | Zendy

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

Author(s) -

Akira Nozawa,

Hideaki Nanamiya,

Takuji Miyata,

Nicole Linka,

Yaeta Endo,

Andreas P.M. Weber,

Yuzuru Tozawa

Publication year - 2007

Publication title -

plant and cell physiology

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 1.975

H-Index - 152

eISSN - 1471-9053

pISSN - 0032-0781

DOI - 10.1093/pcp/pcm150

Subject(s) - transporter , translation (biology) , biochemistry , arabidopsis thaliana , membrane protein , liposome , atp binding cassette transporter , membrane transport protein , chemistry , plant cell , biology , membrane , gene , messenger rna , mutant

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.

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