Fig. 3.

Evidence that the plant COG0212 protein is chloroplast-targeted. a Transient expression in A. thaliana mesophyll protoplasts of green fluorescent protein (GFP) fused to the C terminus of full-length A. thaliana COG0212 (upper panels) or to the predicted targeting peptide of A. thaliana COG0212 (middle panels) and GFP alone (lower panels). GFP (green pseudo-color) and chlorophyll (red pseudo-color) fluorescence were observed by confocal microscopy. Scale bars=10 μm. b Protein import into isolated pea chloroplasts and mitochondria. The full-length A. thaliana COG0212 sequence was translated in vitro in the presence of [³H]leucine. The translation products were incubated for 15 min in the light with chloroplasts (CP) and mitochondria (MT), which were then re-purified on an 8% (v/v) Percoll gradient, without or with prior thermolysin (TH) treatment to remove adsorbed proteins. Proteins were separated by SDS-PAGE and visualized by fluorography. Samples were loaded on the basis of equal chlorophyll or mitochondrial protein content next to an aliquot of the translation product (TP). The positions of molecular mass standards (kilodaltons) are indicated