Fig. 6: Design of Gln-based single α-helical peptides that interact with RAP74-CTD.
From: A glutamine-based single α-helix scaffold to target globular proteins
a Top: design of peptide δ, blending the consensus RAP74-CTD binding motifs of FCP1 with peptide (P3-7)3. Peptide δStpl features a chemical staple covalently linking the side chains at positions 13 and 17, whereas all Leu residues outside the binding motif were substituted by Ala. Peptide δctrl is devoid of helix-stabilizing interactions. Bottom: CD spectra of peptides δ (gold), δStpl (purple), and δctrl (dark red) at 298 K (solid) and 278 K (dashed). b Averaged chemical shift perturbations observed in the 1H-15N resonances of the RAP74-CTD upon the addition of 15 molar equivalents of the (P3-7)3 (teal), δ (gold), δStpl (purple), and δctrl (dark red) peptides. NMR-derived KD values (Supplementary Figure 11a) are shown in the legend. The dashed line sets the threshold for the top 10% peaks with the most intense averaged CSPs upon the addition of δ. c Left: structure of the RAP74-CTD domain of TFIIF (PDB entry 1NHA) colored in a gradient representing the averaged CSPs observed upon the addition of 15 molar equivalents of the δ peptide (white, minimum CSPav; gold, maximum CSPav). Right: region of the overlaid 1H-15N BEST-TROSY spectra of RAP74-CTD in the absence (black) and in the presence of 15 molar equivalents of the (P3-7)3 (teal), δ (gold), δStpl (purple), and δctrl (dark red) peptides. d Top: design of peptide γ blending the RAP74-CTD binding motif AR-WHTLF with peptide (P3-7)2. Peptide γStpl features a chemical staple covalently linking the side chains at positions 10 and 14, whereas all Leu residues outside the binding motif were substituted by Ala. Peptide γCtrl is devoid of helix-stabilizing interactions. Bottom: CD spectra of peptides AR-WHTLF (green), γ (blue), γStpl (purple), and γCtrl (dark red) at 298 K (solid) and 278 K (dashed). e as in b for (P3-7)2 (dark gray), AR-WHTLF (green), γ (blue), γStpl (purple), and γCtrl (dark red). f Left: as in c, left, for peptide γ. Right: as in c, right, for the (P3-7)2 (purple), AR-WHTLF (green), γ (blue), γStpl (purple), and γCtrl (dark red) peptides.
