Figure 2.

Pex3p is a peroxisomal integral membrane protein exposed to the cytosol. (A) Pex3p is localized to peroxisomes. Wild-type E122 cells incubated in oleic acid-containing YPBO medium for 8 h were processed for immunofluorescence microscopy with guinea pig primary antibodies to Pex3p and rabbit primary antibodies to the PTS1 tripeptide SKL. Primary antibodies were detected as described in the legend to Figure 1A. Bar, 2.5 μm. (B) Pex3p exhibits properties of an integral membrane protein. A functionally complementing form of Pex3p tagged with the c-Myc epitope (MycPex3p) resists extraction from a 20KgP subcellular fraction by treatment with 1.0 M NaCl, 1.0 M urea, and 0.1 M Na₂CO₃, pH 11. (C) Pex3p is exposed to the cytosol. MycPex3p in a 20KgP subcellular fraction is sensitive to exogenously added trypsin in both the absence and the presence of the detergent Triton X-100. In contrast, the peroxisomal matrix protein thiolase (THI) is resistant to exogenously added trypsin in the absence of detergent and sensitive in the presence of detergent.