Figure 4. DM binds with fast kinetics to DR-peptide complexes without engaged peptide N-terminus.
(a, b) The hydrogen bonding network between DR1 and full length, covalently linked HA306-318 peptide (a) is compared to a linked HA mutant peptide lacking three N-terminal residues (b). Peptide residues are numbered from P-2 to P11. DRα Val65 was mutated to cysteine to enable disulfide bond formation to HA peptides with a cysteine at the P6 position (HA6 peptides). (c) Space filling model of the empty DR1 groove showing residues contacted by two N-terminal peptide residues (red) and the P1 anchor (green). (d, f) DM binding was enabled by loss of two N-terminal residues and the P1 anchor. DR1 complexes with linked HA6 peptides (5 μM) were injected for 2 minutes (pH 5.35, 15 μl/min 25 °C), followed by injection of buffer. (e) C-terminal truncation of HA6 peptide did not result in substantial DM binding. Complexes linked at the P6 position were tested as in (d). Data are representative of at least three (d, f) and of two (e) independent experiments.