FIGURE 1.

NMR conformational characterization of the T1–898(G901S) peptide. A, the sequence diagram of the peptide shows the characteristic 1-residue stagger. Leading, middle, or trailing chain stagger assignment is indicated as chain 1, 2, or 3, respectively. The ¹³C/¹⁵N doubly labeled residues are underlined and colored in blue, and the Gly to Ser substation site is shown in red. B, the HSQC spectrum of the T1–898(G901S) peptide at 15 °C. The peaks corresponding to the monomer and trimer state are denoted with a superscript M or T, respectively. Chain stagger is indicated as a number in front of the superscript T. C, comparison of experimental NOEs of T1–898(G901S) peptide with a standard triple helical conformation. A predicted contact map obtained from a standard triple helix model structure is shown in shaded boxes as background (14). Contacts are shaded in gray for intrachain distances less than 5 Å and in yellow for interchain distances less than 5 Å. Experimental NOEs for T1–898(G901S) peptide are represented by circles (HN-HN (green circles), HN-H^α (red circles), and HN side chain protons (blue circles)) and are overlaid on the shaded contacts, showing the expected intrachain and interchain NOEs and one new contact (¹Ser¹⁶ NH to ³Ser¹⁶ NH) consistent with the 1-residue stagger between chains and the packing of the Ser residues at the substitution site. Half circles represent overlapped NOEs. The superimposition of the experimental NOEs and the background indicates the 1-residue stagger of the triple helix throughout the substitution site. The diagnostic interchain NH–NH and NH–H^α NOEs between the three chains of the Gly¹³ residues (¹Gly¹³NH to ²Gly¹³NH and ²Gly¹³H^α, ²Gly¹³NH to ¹Gly¹³NH and ¹Gly¹³H^α, ²Gly¹³NH to ³Gly¹³NH and ³Gly¹³H^α, ³Gly¹³NH to ²Gly¹³NH) and between Gly¹⁹ residues (³Gly¹⁹NH to ²Gly¹⁹H^α) are seen as well as interchain NOEs from ²Gly¹⁹NH to ³Pro¹⁷H^β/γ supporting the 1-residue stagger. In addition, the NOEs normally unique between Gly are observed between the Ser¹⁶ residues and residues Gly¹³ and Gly¹⁹ (¹Ser¹⁶NH to ³Gly¹³NH, ³Ser¹⁶NH to ¹Gly¹⁹NH), suggesting that the Ser¹⁶ residues are packed into the center of the triple helix at the substitution site, in a behavior similar to a Gly at the same position. D, the histogram of the ³J_HNHα coupling constants for the T1–898(G901S) peptide. ³J_HNHα coupling constants from the two H^α Gly residues are shown in dark gray and light gray bars.