Figure 3.

Two-dimensional analysis of biotinylated proteins of envelope, thylakoid, and stroma from spinach chloroplast. Chloroplast envelope (E), thylakoid (T), and stroma (S) proteins (200 μg) prepared from biotinylated spinach chloroplasts were passively loaded into 7 cm pH 4–7 IPG strips. Second-dimension separation was in 13% SDS polyacrylamide gels. Proteins were either stained with coomassie blue (A) or transferred to nitrocellulose membrane for biotine detection after Streptavidin Horseradish Peroxidase conjugated analysis (Streptavidin-HRP) (B) or for western-blotting using specific markers of stroma (anti-KARI) or thylakoids (anti-CF1) (C). Major biotinylated spots were indicated on a representative gel (C). In the case of spot 9, a longer exposure of the nitrocellulose membrane, revealing biotinylation is also shown. Rbc SSU, Rubisco small subunit; Rbc LSU, Rubisco large subunit; oec, oxygen evolving complex; plas, plastocyanin. Apparent molecular weight (Mr) in kDa are indicated on the left.