The targeting of the atToc159 preprotein receptor to the chloroplast outer membrane is mediated by its GTPase domain and is regulated by GTP.
The multimeric translocon at the outer envelope membrane of chloroplasts (Toc) initiates the recognition and import of nuclear-encoded preproteins into chloroplasts. Two Toc GTPases, Toc159 and Toc33/34, mediate preprotein recognition and regulate preprotein translocation. Although these two proteins account for the requirement of GTP hydrolysis for import, the functional significance of GTP binding and hydrolysis by either GTPase has not been defined. A recent study indicates that Toc159 is equally distributed between a soluble cytoplasmic form and a membrane-inserted form, raising the possibility that it might cycle between the cytoplasm and chloroplast as a soluble preprotein receptor. In the present study, we examined the mechanism of targeting and insertion of the Arabidopsis thaliana orthologue of Toc159, atToc159, to chloroplasts. Targeting of atToc159 to the outer envelope membrane is strictly dependent only on guanine nucleotides. Although GTP is not required for initial binding, the productive insertion and assembly of atToc159 into the Toc complex requires its intrinsic GTPase activity. Targeting is mediated by direct binding between the GTPase domain of atToc159 and the homologous GTPase domain of atToc33, the Arabidopsis Toc33/34 orthologue. Our findings demonstrate a role for the coordinate action of the Toc GTPases in assembly of the functional Toc complex at the chloroplast outer envelope membrane.
Smith
Matthew D
MD
Department of Biochemistry and Molecular Biology, University of Massachusetts, Amherst, MA 01003, USA.
Hiltbrunner
Andreas
A
Kessler
Felix
F
Schnell
Danny J
DJ
D020816:Amino Acid Motifs; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002736:Chloroplasts / Q000235:genetics / Q000378:metabolism*; D016384:Consensus Sequence; D004926:Escherichia coli / Q000235:genetics; D020558:GTP Phosphohydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism*; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017354:Point Mutation; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000737:chemistry* / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000502:physiology; D013820:Thermolysin / Q000494:pharmacology
D029681:Arabidopsis Proteins; D008565:Membrane Proteins; D011498:Protein Precursors; C403444:TOC159 protein, Arabidopsis; D006160:Guanosine Triphosphate; D013820:Thermolysin; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins