Xenbase Image ID: 156420

Figure EV5. XPF leucine 219 is part of the major interaction site between XPF and SLX4 XPF‐ERCC1‐depleted (ΔXE) and XPF‐ERCC1‐depleted NPE supplemented with XPF‐ERCC1 (+XEWT), XPF‐ERCC1 and SLX4 (+SXEWT), or XPFL219R‐ERCC1 and SLX4 (+SXEL219R) were analyzed by Western blot using α‐XPF and α‐SLX4 antibodies. Extracts were used for Fig 6A. As in (A). Replicate of Fig 6A. The extracts from (B), with similarly treated HSS, were used to replicate pICL. Samples were taken at the indicated times and analyzed by XPF (left) and SLX4 (right) ChIP using pICL and pQuant primers. The qPCR data were plotted as the percentage of peak value with the highest value set to 100%. Replicate of Fig 6C. Wild‐type and mutant FLAG‐XPF‐ERCC1 were co‐expressed with His‐SLX4 in Sf9 insect cells. Cells were lysed and XPF was immunoprecipitated via the FLAG‐tag. Samples were analyzed by Western blot using α‐FLAG and α‐His antibodies. In, input; FT, flow‐through fraction; B, fraction bound to beads. Size exclusion chromatography of recombinant XPF‐ERCC1 and BTB domain of SLX4. Superdex 200 gel filtration column elution profile of FLAG‐XPF‐ERCC1, His‐tagged BTB domain, and both proteins combined (top panel). The XPF‐ERCC1 heterodimer eluted at ˜12 ml, while His‐BTB eluted around ˜16 ml. Collected fractions during elution were analyzed by Western blot using α‐XPF and α‐His antibodies (bottom panel). The BTB domain protein does not shift to a higher elution volume when incubated with XPF‐ERCC1 indicating the affinity is not high enough to show binding between the two proteins. Source data are available online for this figure. Image published in: Klein Douwel D et al. (2017) © 2017 The Authors. This image is reproduced with permission of the journal and the copyright holder. This is an open-access article distributed under the terms of the Creative Commons Attribution license Larger Image Printer Friendly View

Return to previous page