Binding of synthetic peptides by a human monoclonal IgM with an unusual combining site structure.
Using X-ray crystallography, a human monoclonal IgM cryoglobulin (Mez) was found to have an unusual combining site topography. Analysis of the unliganded Fv at 2.6 A resolution revealed that the HCDR3 had partitioned the active site into two compartments [Ramsland PA et al. 2000. Mol. Immunol. 37: 295-310]. The two cavities had dimensions and chemical properties that were compatible with the binding of peptides. In this study, libraries of peptides were prepared using solid-phase synthesis. Binding of the intact Mez IgM to these peptides was tested by enzyme-linked immunoassays. Screening of 400 dipeptides revealed that binding was markedly skewed toward amino acids with aromatic side-chains (Phe and Trp), especially when located in the second position. Preferential recognition of aromatic side-chains by Mez IgM was confirmed with larger peptides of three to five residues, but C-terminal positioning was not favored in these peptides. Mez IgM also showed binding propensities for acidic residues (Asp and Glu) as well as several other side-chains with different chemical properties, including His, Pro, Asn and Gln. Mez IgM recognized sets of overlapping octapeptides representing the sequences of the constant domains of human IgG1 heavy chains. These peptides represented similar stretches of polypeptide on the three-dimensional structures of all three constant domains (CH1, CH2 and CH3). Thus, Mez IgM may recognize structurally homologous regions of immunoglobulin domains, which were conserved during the evolution of the immune system.
Edmundson
A B
AB
Crystallography Program, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA. Allen-Edmundson@omrf.ouhsc.edu
Tribbick
G
G
Plompen
S
S
Geysen
H M
HM
Yuriev
E
E
Ramsland
P A
PA
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology; D000915:Antibody Affinity; D001666:Binding Sites, Antibody / Q000276:immunology; D020650:Combinatorial Chemistry Techniques; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D006801:Humans; D007075:Immunoglobulin M / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008024:Ligands; D008969:Molecular Sequence Data; D019151:Peptide Library; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D008258:Waldenstrom Macroglobulinemia / Q000276:immunology
D000911:Antibodies, Monoclonal; D007075:Immunoglobulin M; D008024:Ligands; D019151:Peptide Library; D010455:Peptides