Modulation of the Peptide-Binding Specificity of a Single-Chain Class II Major Histocompatibility Complex

抄録

We designed and expressed a single-chain class II major histocompatibility complex molecule capable of forming a stable complex with an antigenic peptide. The peptide-binding preference of the single-chain (sc) human leukocyte antigen derived from DRB5'0101 (DR 51) was determined to be similar to that of the authentic one, which requires a bulky hydrophobic residue at position-1 (P 1) as a primary anchor. For modulation of the peptide-binding affinity, we modified binding pocket 1 of sc DR 51 by sitedirected mutagenesis. The relative binding affinity of the engineered sc DR 51 for several P 1-substituted peptides was measured by competition assaying with a fluorescence labeled peptide. The se DR 51 molecule showed high affinity to the self-peptide derived from myelin basic protein, 87-98 with Phe as the P1 residue (F 90 F). While reduction of pocket 1 volume (β G 86 V) decreased the affinity of F 90 F, it rather increased the affinity of the Ala-substituted peptide as to the P 1 residue (F 90 A). Through more extensive engineering in the peptide-binding groove of the sc DR 51 molecule, it is expected that we can construct sc DR 51 variants with various peptide ligand motifs.