Fig. 5

Membrane orientation of PDI8. a Immunoblot analyses of proteins extracted from 35S _pro :PDI8 plants. The 35S _pro :PDI8 total protein homogenate was separated into soluble (sol) and microsomal membrane (mem) fractions by centrifugation. Immunoblots were incubated with affinity-purified anti-PDI8 antiserum (upper panel). A polyclonal goat anti-BiP antibody was used as a marker for microsomes (middle panel). The large subunit of Rubisco (LSU) was used as a marker for the soluble phase in an SDS-PAGE gel stained with Coomassie (lower panel). b Protease protection assays were performed with 35S _pro :PDI8 microsomes. Samples were either treated (+) or not treated (-) with 50 μg/mL proteinase K (PK) and 0.1 % Triton X-100 (TX), and immunoblot analysis was performed using the anti-PDI8 antiserum. c Model of the PDI8 polypeptide oriented in the ER membrane