JCS008367 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig. S1. Relationship between the apparent diffusion constant and the molecular mass of a protein. The mobility of EGFP alone (30 kDa) and of the EGFP-tagged NER proteins XPA (70 kDa), XPG (210 kDa), ERCC1-XPF (215 kDa) and TFIIH (500 kDa) was determined by FRAP and subsequent Monte Carlo simulations, yielding apparent diffusion constants for all fusion proteins and EGFP alone. The data points were fitted with the exponential equation y=246x⁻⁰,⁷³, where x is the molecular mass in kDa and y the apparent diffusion constant in μm²/second. Using this equation, diffusion constants were derived for monomeric DDB2 (78 kDa), dimeric UV-DDB (205 kDa), multi-protein complex DDB-CUL4A-ROC1 (310 kDa) and DDB-CUL4A-ROC1 associated with COP9 (760 kDa). The calculated diffusion constants are plotted in the same graph as the measured diffusion constant for DDB2-EYFP that fits well to the expected diffusion constant of DDB-CUL4A-ROC1 associated with COP9. All values include an additional 30 kDa because of the presence of yellow or green fluorescent protein.

• Supplemental Figure S2 -

  Fig. S2. Accumulation of endogenous DDB2 and TFIIH (p89 subunit) after local UV damage (30 J/m²) in wild-type human cells (VH10) and several NER mutant cell-lines (XP-C, XP-A, XP-G and XP-F). The left column show the immunofluorescent labeling with anti-XPB antibody (green) and the right column shows labeling with anti-DDB2 antibody (red).