Mouse Immunoglobulin Subclasses: Cyanogen Bromide Fragments and Partial Sequence of a gamma1 Chain

Abstract

The purification and characterisation of all the cyanogen bromide fragments of MOPC 21 heavy (yl) chain is described. The ten BrCN fragments account for the whole chain. Four of these fragments have been used to establish the sequence of the C-terminal stretch (138 residues) that includes the entire CH3 and a part of the CH2 homology regions. A comparison of this sequence with homologous sequences is presented. Mouse y l and y2 proteins differ much more than the y subclasses in humans and in other species. The comparison further suggests that the four human y subclasses and mouse y l have a common ancestor which differs from the mouse y 2 ancestor. Unlike other subclasses, mouse y l and y 2 genes have diverged before speciation.Heavy and light chains of immunoglobulins contain an amino-terminal variable portion (the V-region) of 100 -120 residues, and a carboxy-terminal "constant" portion (the C-region) which includes the remainder of the chain. All the heavy chains share a common pool of V-region genes while a cluster of C-region tandem genes defines the variety of classes and subclasses of heavy chains. (For a recent review, see [l] and [2].)Comparative sequence studies, especially of human IgG subclasses, led to the suggestion that subclasses resulted from recent tandem gene duplications and that the genes were under a constant process of expansion and contraction [3]. Clearly, more extensive studies were required to test the generality of these observations. Mouse immunoglobulins are particularly useful for such studies because of the availability of myeloma proteins [4]. Murine immunoglobulins include three major immunoglobulin G subclasses which differ in their interchain bridges [5,6]. Sequences around their intrachain bridges are also available [6,5], and extensive sequence studies on a murine y2a protein (MOPC 173 H-chain) have been reportedThe work described here was done on the mouse plasmacytoma MOPC 21 which contains a y l (heavy) chain. Specific interest in this chain derives from the [7,81.

Sequence analysis of cloned cDNA encoding part of an immunoglobulin heavy chain

“…Representative 'ladders' are shown in Figure 3, and the complete DNA sequence is in Figure 4. It covers 459 nucleotides between the A.T joints, and encodes amino acids 287 to 439 in the sequence of Adetugbo (15,16). This includes half of the C H2 domain (amino acids 228-334)…”

“…The individual nucleotides in the sequences of the A.T joints were only partially resolved, but measurements of the complete poly-A blocks on ladders indicated lengths of 76 +15 bp on the left and 50 +15 bp on the right. The The DNA sequence implies several substitutions in the published amino acid sequence (15,16), as listed in Table I. In all but the first, the DNA sequence appears certain.…”

“…Other amino acid sequences are from refs. (15,25). AA = amino acid; + = insertion; * = DNA sequence uncertain.…”

“…Nucleic Acids Research although not in a p chain (15,(23)(24)(25)(26)(27)(28). Our DNA sequence is uncertain at that point but appears not to encode valine.…”

“…(3) Protein sequencing errors. 4 of the 8 substitutions involve the interchange of pairs of nearby amino acids; one involves an acid-to-amide change; and one (codon 377) would replace a serine which was not definitely present in the peptide sequenced (15) with a tryptophan which could have been present in greater molarity than estimated (15).…”