No abstract

The antibody-binding site is formed as a result of the association between VH and VL domains. Several studies have shown that this association plays an important role in the mechanism of antigen-antibody interaction (Stanfield et al. Structure 1: 83-93, 1993). Considering this, we propose that variations in the VH-VL association are part of the diversification strategy of the antibody repertoires. Previously, a model of association for VH-VL domains based on geometrical characteristics of the packing at the interface was developed by Chothia et al. (J. Mol. Biol. 186: 61-663, 1985). This model includes a common association form for antibodies and a three-layer structure for the interface. In the present work, a complementary model is introduced to account for the general geometrical restrictions of the VH-VL interface, and particular arrangements related to the chemical properties or the side-chain orientations of participating residues. Groups of residues assume common side-chain orientations, which are apparently related to particular functions of different interface zones. Analyses of amino acid usage and network are in agreement with the side-chain orientation patterns. Based on these observations, a three-zone model has evolved to illuminate geometrical and functional restrictions acting over the VH-VL interface. Additionally, this study has revealed the asymmetrical relationships between VH and VL residues important for the association of the two domains.

Although human and mouse antibodies are similar when one considers their diversification strategies, they differ in the extent to which kappa and lambda light chains are present in their respective variable light chain repertoires. While the Igk-V germline genes are preponderant in mice (95% or more), they comprise only 60% in humans. This may account for differences in the structural repertoire encoded in the Igk-V germline genes of these species. However, this subject has not been properly investigated, partially because a systematic structural characterization of the mouse Igk-V germline genes has not been undertaken. In the present study we compiled all available information on mouse Igk-V germline genes to characterize their structural repertoire. As expected, comparison with the structural repertoire of human Igk-V germline genes indicates differences. The most interesting is that the mouse Igk-V germline gene repertoire is more diverse in structural terms than its human counterpart: the mouse encodes seven canonical structure classes (combination of canonical structures in L1 and L3). In contrast, the human encodes only four. Analysis of the evolutionary relationships of human and mouse Igk-V germline genes led us to propose that the difference reflects a strategy of mice to compensate for the small lambda chain contribution to the repertoire of their variable light chains.

Variable genes of human Ig are classified in families and clans which reflect the early events of gene duplication in the evolution of the locus. This organization in multiple copies of variable genes plus the somatic processes of recombination and hypermutation allows the immune system to generate an antibody repertoire of great diversity. At present the role that somatic processes play in the generation of that diversity is understood with some detail. It is a matter of hard controversy, however, which selective pressures have shaped the evolution of the germline genes of Ig and, consequently, what the role of this germline component in the generation of the antibody diversity actually is. Previous studies of our group have showed that the structural repertoire of Ig-determined by the canonical structures-is an important factor to determine the recognition properties of the antibodies. Complete knowledge of the sequences of the human V(H) and Vkappa loci is available to analyze the evolution of the structural repertoire of these loci. Two phylogenetic gene trees were built from the functional germline genes and the evolution of the structural repertoire was studied. We report that for both loci the canonical structures are not randomly distributed within the tree. Conversely, it is shown that the evolution of the structural repertoire follows a gradual process of diversification. This indicates a correlation between the evolution of genes and the structural repertoire, although important differences are found in the patterns of evolution of the structural repertoire between V(H) and Vkappa. Based on those results we propose a primordial structural repertoire for V(H) and Vkappa. The general properties and an outline of the three-dimensional structure of this primordial repertoire are given.

The antibodies of known three-dimensional structure exhibit a definite number of conformations (canonical structures) for five of six hypervariable loops. In the present study it was found that approximately 85% of the immunoglobulin sequences analyzed fall into a small number of canonical structure combinations, representing only 3% of the total possible. These structures were classified into six distinct groups, depending on the type of antigen with which they interact. Within each loop, the positions responsible for maintaining these canonical structures show a use frequency of amino acids that fits an inverse power law, whereas the use frequency of the amino acids responsible for the detailed antigenic specificity follows an exponential distribution. We propose an evolutionary interpretation that connects these data, using the fact that the inverse power law is generated by statistical processes of the type that yield a wealth curve and the fact that exponential distribution is generated by processes that are not biased by past history.