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. 2010 Oct 29;285(53):41244–41254. doi: 10.1074/jbc.M110.155242

FIGURE 4.

FIGURE 4.

Dbf4 binds a surface on the PBD distinct from its phospho-protein binding site. A, a biotinylated Spc72 phospho-peptide (residues 223–242) bound the PBD in the AlphaScreen assay. B, the same (non-biotinylated) Spc72 phosphopeptide did not compete the Dbf4-PBD interaction. C, purified wild type PBD and PBD-HK proteins interact with Dbf4 in the AlphaScreen assay, but the PBD-HK mutant protein fails to interact with Spc72 phosphopeptide. D, two-hybrid Spc721–400 and Dbf466–227 interactions with the PBD were tested on the indicated plates. As in C, mutation of the PBD pincer residues H641A and K643M or W517F, H641A, and K643M had no affect on the Dbf4-PBD interaction but eliminated the Spc72-PBD interaction. 3-AT, 3-aminotriazole.