The amino acid sequence of the heavy-chain variable region of the crystallizable human myeloma protein Dob has been determined. This protein has previously been shown to have a deletion in the hinge region [Lopes, A. D., & Steiner, L. A. (1973) Fed. Proc., Fed. Am. Soc. Exp. Biol. 32, 1003; Steiner, L. A., & Lopes, A. D. (1979) Biochemistry (preceding paper in this issue)]. The complete sequence was established by analysis, in the automated sequenator, of the intact Fd' piece and of three large overlapping fragments prepared from Fd' by digestion with cyanogen bromide, by tryptic digestion of the citraconylated Fd', and by cleavage with hydroxylamine. Portions of the sequence were confirmed by examination of the amino acid composition and the partial sequence of a variety of small peptides obtained by enzymatic degradation. The Dob heavy-chain variable region appears to belong to the VHIII subgroup, but there are several unusual substitutions. Residue 45 in the Dob sequence is proline, although all other known heavy-chain sequences in man, mouse, rabbit, and guinea pig have leucine at this position. Positions 10 (aspartic acid), 68 (alanine), and 82 (leucine) in the Dob sequence are also atypical. There is no deleted segment in the variable region of the Dob heavy chain nor any abnormality in the variable-constant joining region. The hinge-region deletion appears to be the only gross structural anomaly in the Dob heavy chain.

The partial amino acid sequence of pooled, microheterogeneous pig immunoglobulin lambda chains was determined previously (Franĕk, F. (1970), FEBS Lett. 8, 269; Novotný, J., and Franĕk, F. (1975), FEBS Lett. 58, 24). In the present study, citraconylated pig lambda chains were digested by trypsin under conditions in which some of the epsilon-amino groups of lysine residues unmask. The resulting fragments were purified by gel filtration and ion-exchange chromatography at pH 3.0 in buffers containing urea; some of the fragments were found to be of intermediate size (i.e., larger than normal tryptic peptides but smaller than "citraconyl" peptides), thus permitting overlap information and amino acid sequences of all the 14 tryptic peptides to be deduced from amino acid compositions and partial amino acid sequences of selected fragments. In addition to completing the major amino acid sequence of pig immunoglobulin lambda chains, the present study demonstrates that it is possible to sequence microheterogeneous proteins with a suitable fragmentation strategy.

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