CRYSTALLIZABLE HUMAN MYELOMA PROTEIN DOB HAS A HINGE-REGION DELETION

During experiments to prepare heavy-metal derivatives of the crystallizable human IgGl (k) immunoglobulin Dob, it became apparent that this protein has several unusual features. (1) Instead of the four labile interchain disulfide bridges ordinarily found in IgGl, the Dob protein has only a single interchain disulfide bridge, which connects its two light chains. (2) The Dob heavy chain appears to be slightly smaller than a control γ1 chain, as judged by polyacrylamide gel electrophoresis in sodium dodecyl sulfate and by gel filtration in guanidine. (3) The Dob heavy chain has three fewer residues of half-cystine than expected in γ1 chains. (4) The Dob IgG is relatively resistant to digestion with papain and trypsin; however, it is readily digested with pepsin, although at an unusual site. These findings suggest that some or all of the γ1 hinge region is missing in Dob. To localize the deletion, we prepared an F(ab')2 fragment consisting of two heavy-chain pieces (Fd') noncovalently associated with the light-chain dimer. The Fd' piece was isolated and digested with trypsin. The sequence of the C-terminal tryptic peptide was Val-Ala-Pro-Glu-Leu-Leu-Gly-Gly-Pro-Ser-Val. Positions 2-11 of this peptide are identical with residue positions 231-240 of the γ1 chain. The N-terminal valine could be either Val-211 or Val-215 of the γ1 sequence. A tryptic peptide, Val-Asp-Lys-Lys, was also isolated from Dob Fd'; this sequence is not found in the variable region of the Dob heavy chain [Steiner, L. A., Garcia Pardo, A., & Margolies, M. N. (1979) Biochemistry (following paper in this issue)] but corresponds to positions 211-214 of the γ1 constant region. Therefore, the deletion cannot include these residues and must begin after Val-215; normal γ1 sequence resumes at Ala-231. The same 15-residue deletion has been found in two other IgGl proteins, Meg [Fett, J. W., Deutsch, H. F., & Smithies, O. (1973) Immunochemistry 10, 115] and Lec [Rivat, C., Schiff, C., Rivat, L., Ropartz, C., & Fougereau, M. (1976) Eur. J. Immunol. 6, 545]. Possible explanations for the occurrence of identical hinge-region deletions in three different immunoglobulins are suggested by recent experiments demonstrating that the three constant domains and the hinge region of mouse γ1 chains are each encoded by separate segments of DNA [Sakano, H., Rogers, J. H., Hüppi, K., Brack, C., Traunecker, A., Maki, R„ Wall, R„ & Tonegawa, S. (1979) Nature (London) 277, 627]. © 1979, American Chemical Society. All rights reserved.