Figure 5.

TrJ Pmp22 exists mainly as dimers, not monomers, in Nonidet P-40 buffer. COS7 cells expressing Myc-tagged TrJ Pmp22 were crosslinked with DSP and lysed in Nonidet P-40 buffer. The lysates were separated directly on the gel filtration column in the same buffer (A) or denatured (as described in Materials and Methods) before the chromatography (B). The fractions were treated with the sample buffer without the reducing reagent DTT before being separated by SDS/PAGE. TrJ Pmp22 was visualized by Western blot with anti-c-Myc antibody. In A, both uncrosslinked monomers and crosslinked dimers of TrJ Pmp22 were detected in the dimer peak. However, in B, the uncrosslinked TrJ Pmp22 dimers now appeared in the monomer peak, whereas the crosslinked TrJ Pmp22 dimers still eluted in the dimer peak.