Sequence Relationships among the Variable Regions of Immunoglobulin Heavy Chains from Various Mammalian Species

Abstract

Immunoglobulin heavy chains of myeloma proteins from dogs and cats have been subjected to automated sequence analysis. When the results were compared with human heavy-chain sequences, all the dog and cat proteins could be unequivocally assigned to the VHIII subgroup. This pattern contrasts with that in human proteins in which only 25% of all heavy chains sequenced belong to this subgroup. The 30 residues at the NH2 termini of dog, cat, and human heavy chains had sequence identities near or exceeding 90%, in contrast to established interspecies sequence homologies of constant regions of about 60%. Some genes of the immunoglobulin heavy-chain variable region thus appear to have been conserved through a considerable period of evolutionary time.The analyses also showed that the presence of certain amino acids at certain positions in these heavy chains could be correlated with the species of origin. The occurrence of such "phylogenetically associated" residues is most consistent with the presence of a restricted number of genes in the heavy-chain variable region pool.Both heavy (H) and light (L) polypeptide chains of immunoglobulins contain two distinct regions from the standpoint of variability of amino-acid sequence (1, 2). The variable (V) region is found in the amino-terminal portion of each chain and is believed to participate in the antigen-binding function of these molecules. The constant (C) region comprises the balance of each chain and, in the heavy chain, mediates the biological, or effector, properties characteristic of many immunoglobulins (3). Several lines of evidence suggest that the variable and constant regions of a given H or L chain are encoded by separate genes (4, 5).The pattern of sequence variability among variable regions is such that certain collections of individual sequences are much more closely related to each other than to other variable region sequences. These ensembles of related sequences, first noted for light chains (6-8), are referred to as variable region subgroups and are found in both kappa and lambda light chains. More recently, four subgroups have been identified in heavy chains (9)(10)(11)(12). Of the four, only the VHIII subgroup has an unblocked (i.e., not pyrollidone carboxylic acid) aminoterminal residue.Sequence analyses of heavy-chain variable regions of an unselected series of homogeneous myeloma proteins isolated from dogs and cats are reported here. A comparison of these sequences with certain human proteins suggests an unanticipated degree of evolutionary stability for certain genes of the immunoglobulin variable region. MATERIALS AND METHODSThe source and methods of preparation of the various myeloma proteins have been reported elsewhere (13,14). IgG fractions were prepared from pooled sera by DEAESephadex ion-exchange chromotography. Purified heavy chains were subjected to direct sequence analysis on a Beckman model 890A automated sequencer (15). RESULTS AND DISCUSSIONThe relevant variable region sequences are shown in Fig. 1. The canine and feline heavy ...

Phylogenetically Associated Residues Within the Vhiii Subgroup of Several Mammalian Species

“…The heavy chain result in the opossum (a pouched mammal) assumed particular interest because, although the sequence of the first 11 residues of the pool showed unequivocally that this preparation was assignable to the Vmii subgroup, the residue at position 2 was an isoleucine rather than the valine that is characteristic of VlcIIII proteins (30,13). This undoubtedly represents a "phylogenetically associated" residue (10) and merits further analysis in other members of the Metatheria.…”

“…For example, position 11 in rabbit kappa chains from homogeneous antibodies has generally been valine, while mouse and human kappa chains contain leucine at this position. The identification and interpretation of such residues have been complicated by the difficulty of specifying precisely what is implied by the terms "species specific" or "phylogenetically associated" (10). The residues in question a~e rarely, if ever, unique to a given species but are more accurately associated with broader level phylogenetic groups.…”

“…4 compares the gas chromatographic analysis of position 23 of representative canine and murine myeloma proteins with pools from these same species. Previous studies (10) showed that all of a series of canine and feline myeloma proteins had valine in position 23 while human and murine myelomas had alanine here. A comparison of the data from the pooled heavy chains from these species shows that the canine pool contains exclusively valine at this position while the murine pool contains exclusively alanine (Fig.…”

“…Previous studies of heavy chains prepared from myeloma proteins have shown that the primary structure of the variable region of proteins in the V n m subgroup is extremely conserved in at ]east four species, and that phylogenetically associated residues are identifiable in such proteins (10). The knowledge that the VHm subgroup is the only unblocked heavy chain subgroup (11-13) suggested that it might be possible to search for such phylogenetically associated residues in pooled heavy chains from a number of mammalian species.…”

Amino-Acid Sequence of the Variable Region of the Heavy (Alpha) Chain of a Mouse Myeloma Protein with Anti-Hapten Activity

“…Each has an unblocked aminoterminal (Glu or Asp), which is characteristic of human VH sequences of the VHIII subgroup (21)(22)(23), and it has been suggested that this subgroup may be the predominant one in some nonprimate species (24). However, the sequences of VH173 and VH315 differ markedly; the two are only 40% homologous and they differ by at least two sequence gaps (Fig.…”