Purified Sym and Ssu72 were incubated and isolated by preparative gel filtration.

Interaction in vitro; bait produced as a recombinant fusion protein in bacterial system; prey produced as a recombinant fusion protein in bacterial system.

Fig3, Tbl1.

Interaction in vitro; protein produced as a recombinant fusion protein in bacterial system; peptide commercially synthesized.

In this experiment, Ssu72 is complexed with Sym to increase its stability and is the catalytically inactive C13D/D144N form. The crystal structure shows the best fit with a RpII215 peptide phosphorylated on Ser5 explaining preference for this substrate.