Figure 4.

Recognition of the NIM Peptide by Nrd1p CID

(A) Solution structure of Nrd1p CID bound to the NIM peptide. The NIM peptide is represented in yellow sticks (only nonhydrogen atoms are shown), and the protein is shown as a gray ribbon model. Protein residues that form hydrophobic contacts and putative hydrogen bonds to the NIM peptide are shown in magenta sticks.

(B) Electrostatic surface representation of Nrd1p CID (electropositive in blue, electronegative in red, neutral in white) with the NIM peptide (represented in yellow sticks; only nonhydrogen atoms are shown). The upstream electronegative stretch of NIM interacts with the electropositive pocket of Nrd1p CID, while the β turn conformation formed by Asn580-Pro581-Tyr582-Thr583 docks in a hydrophobic pocket of Nrd1p CID.

(C) Superposition of Nrd1p CID-Ser5P CTD (blue), Nrd1p CID-Trf4p NIM (yellow), and Pcf11p CID-Ser2P CTD (magenta) complexes, displaying only peptide ribbons on the surface of Nrd1p CID. The comparison highlights the β turn conformation recognition of CTDs and NIM by the CIDs.

(D) Solution structure of Nrd1p CID bound to the Ser5P CTD peptide. The phospho-CTD peptide is represented in yellow sticks (only nonhydrogen atoms are shown), and the protein is shown as a gray ribbon model. Protein residues that form hydrophobic contacts and putative hydrogen bonds to the phospho-CTD peptide are shown in gray sticks.

(E) Scheme showing contacts and energetics between the NIM peptide and Nrd1 CID. Equilibrium binding experiments with both the protein and peptide mutants (in red and blue, respectively) were monitored by FA (for the binding isotherms, see Figure S5). Other residues involved in the canonical CTD-CID interface were mutated previously (Kubicek et al., 2012, Vasiljeva et al., 2008). L20D mutant disrupts the hydrophobic contact with Phe17 and impairs the overall geometry of the α1-α2 loop that contributes to the interaction with the upstream electronegative stretch of NIM.