Figure 2.
Mutagenesis targets within the pre-TCR β subunit. A, pre-TCR crystal structure model (Protein Data Bank code 3OF6) in schematic representation in the same orientation as Fig. 1B. B, expanded view of boxed area of A showing only TCRβ. The view is rotated ∼15° about x and y to highlight the contact surface. Side chains of residues contacting (i.e. within 4 Å of) pTα are shown in stick representation. Three hydrophobic residues targeted for mutagenesis are colored yellow. Numbering is according to that found in the crystal structure 3OF6 and is offset from those of N15 and N30β by three or two residues, respectively. C and D, overlay of 1H-15N TROSY-HSQC of N30β (18) and N30β-c1 (9) (C) or N15β (18) and N15β-c1 (9) (D) to illustrate similarity of spectra. E, region of overlaid spectra shown in C with labeled peaks indicating lack of chemical shift changes in the V domain (Thr-6, Thr-32, Asp-25, Leu-111, and Leu-117), whereas those in the C domain (Val-125, Tyr-184, Arg-189, Gln-209, and Phe-210) show significant changes. F, rotational correlation times of WT and mutant TCRβs suggest abrogation of self-association within Cβ. The plot shows τc versus concentration for N15 (blue diamonds) and N30β (red triangles). Lines show exponential fits of points to guide the eye. WT is shown with filled symbols and solid lines, and c1 mutants are shown with open symbols and dotted lines.