A single-domain antibody fragment in complex with RNase A: non-canonical loop structures and nanomolar affinity using two CDR loops.

TitleA single-domain antibody fragment in complex with RNase A: non-canonical loop structures and nanomolar affinity using two CDR loops.
Publication TypeJournal Article
Year of Publication1999
AuthorsDecanniere, K., A. Desmyter, M. Lauwereys, M. A. Ghahroudi, S. Muyldermans, and L. Wyns
JournalStructure
Volume7
Issue4
Pagination361-70
Date Published1999 Apr 15
ISSN0969-2126
KeywordsAmino Acid Sequence, Animals, Antibody Affinity, Antibody Specificity, Antigen-Antibody Complex, Antigen-Antibody Reactions, Binding Sites, Antibody, Camels, Cattle, Crystallography, X-Ray, Humans, Immunoglobulin Heavy Chains, Mice, Models, Molecular, Molecular Sequence Data, Pancreas, Protein Conformation, Ribonuclease, Pancreatic, Software, Species Specificity
Abstract

BACKGROUND: Camelid serum contains a large fraction of functional heavy-chain antibodies - homodimers of heavy chains without light chains. The variable domains of these heavy-chain antibodies (VHH) have a long complementarity determining region 3 (CDR3) loop that compensates for the absence of the antigen-binding loops of the variable light chains (VL). In the case of the VHH fragment cAb-Lys3, part of the 24 amino acid long CDR3 loop protrudes from the antigen-binding surface and inserts into the active-site cleft of its antigen, rendering cAb-Lys3 a competitive enzyme inhibitor.RESULTS: A dromedary VHH with specificity for bovine RNase A, cAb-RN05, has a short CDR3 loop of 12 amino acids and is not a competitive enzyme inhibitor. The structure of the cAb-RN05-RNase A complex has been solved at 2.8 A. The VHH scaffold architecture is close to that of a human VH (variable heavy chain). The structure of the antigen-binding hypervariable 1 loop (H1) of both cAb-RN05 and cAb-Lys3 differ from the known canonical structures; in addition these H1 loops resemble each other. The CDR3 provides an antigen-binding surface and shields the face of the domain that interacts with VL in conventional antibodies.CONCLUSIONS: VHHs adopt the common immunoglobulin fold of variable domains, but the antigen-binding loops deviate from the predicted canonical structure. We define a new canonical structure for the H1 loop of immunoglobulins, with cAb-RN05 and cAb-Lys3 as reference structures. This new loop structure might also occur in human or mouse VH domains. Surprisingly, only two loops are involved in antigen recognition; the CDR2 does not participate. Nevertheless, the antigen binding occurs with nanomolar affinities because of a preferential usage of mainchain atoms for antigen interaction.

Alternate JournalStructure
PubMed ID10196124