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Figure 2 | BMC Molecular Biology

Figure 2

From: The Drosophila methyl-DNA binding protein MBD2/3 interacts with the NuRD complex via p55 and MI-2

Figure 2

Interactions between MBD2/3 and NuRD homologues in a GST-pulldown assay (A) Analysis of interactions between radioactively labelled SV40 large T antigen (large T) and a number of control GST fusion proteins under stringent buffer conditions. Significant amounts of SV40 large T were only precipitated by p53. The faint bands seen with some of the other proteins were considered background. (B) GST-MBD2/3 fusion proteins for long and small isoforms (MBD2/3li, MBD2/3si, respectively) efficiently precipitated radioactively labelled MBD2/3 long isoform and p55 (solid arrowhead). Similarly, MI-2 protein could be precipitated by the small GST-MBD2/3 isoform (solid arrowhead). No interaction could be observed between long MBD2/3 or small MBD2/3 isoforms and MTA-like or RPD3 (open arrowheads). (C) A MBD2/3-specific antibody immunoprecipitates p55, but not GAGA factor from embryonic nuclear extracts. No proteins were detectable in control immunoprecipitations with a myc-specific antibody. (D) Size-fractionation of baculovirus-expressed MBD2/3 long isoform and RPD3. Baculovirus-expressed MBD2/3li elutes in fractions that significantly exceed the calculated molecular weight of the protein (36 kDa), thus indicating that MBD2/3li efficiently multimerizes in solution. This effect appeared to be specific for MBD2/3 and was not observed with baculovirus-expressed RPD3 protein (58kDa). The size of marker proteins is shown left and on top, IN indicates input protein.

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