New ligands for HLA DRB1*0301 by random selection of favourable amino acids ranked by competition studies with undecapeptide amide sublibraries.
An efficient screening procedure for the identification of high affinity HLA class II ligands and their binding pattern has been established to characterize peptide specificities for the HLA allele DRB1*0301. The method is based on the screening of 209 synthetic undecapeptide amide sublibraries O/X10-NH2 representing collections of 19(10) individual peptides in a competition ELISA using HLA DRB1*0301 protein and the biotinylated natural ligand ApoB 2877-2894. Screening results represent the effect on competition induced by an individual amino acid residue in its sequence position of undecapeptide amides. Amino acids clustered as active in their position were randomly selected for the same position of a restricted set of 96 individual undecapeptide amides. This novel approach for the design of ligands was introduced to compensate for the inaccuracy induced by the translational invariance of amino acids in peptide libraries characterized by one defined amino acid. Translational invariance is facilitated by shifted docking of O/X10-NH2 libraries in the binding cleft and protrusion from the ends of the cleft. A second more directed deduced set of 24 peptides was obtained by combination of the most favourable residues in each position. All individual peptides were investigated in the competition assay. The most active HLA DRB1*0301 ligands were obtained by random selection of favourable amino acids and six of them showed improved affinity in comparison to the model ligand alpha AChR 310-325.
Jung
C
C
Neurologische Klinik der Universitätsklinik Tübingen, Germany.
Kalbus
M
M
Fleckenstein
B
B
Melms
A
A
Jung
G
G
Wiesmüller
K H
KH
D000483:Alleles; D000577:Amides / Q000378:metabolism*; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis* / Q000378:metabolism; D001055:Apolipoproteins B / Q000378:metabolism; D001665:Binding Sites; D001667:Binding, Competitive; D019921:Biotinylation; D016208:Databases, Factual; D004797:Enzyme-Linked Immunosorbent Assay; D006684:HLA-DR Antigens / Q000235:genetics / Q000378:metabolism*; D059811:HLA-DRB1 Chains; D006801:Humans; D020128:Inhibitory Concentration 50; D020407:Internet; D008024:Ligands; D008954:Models, Biological; D009842:Oligopeptides / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D019151:Peptide Library; D016415:Sequence Alignment
D000577:Amides; D000596:Amino Acids; D001055:Apolipoproteins B; D006684:HLA-DR Antigens; D059811:HLA-DRB1 Chains; C553502:HLA-DRB1*03:01 antigen; D008024:Ligands; D009842:Oligopeptides; D010446:Peptide Fragments; D019151:Peptide Library