Continuation of a comprehensive program is proposed aimed at the comparative structural study of the pathological IgG, IgA, IgM, and IgD immunoglobulins present in large amount in the plasma of patients with the lymphocytic diseases, Waldenstrom's macroglobulinemia and multiple myeloma, respectively. The overall objectives are: 1) to determine the complete covalent structure of IgM and IgD immunoglobulins from myeloma patients, 2) to relate these structures to subgroup, subclass, class, and biological differences among human immunoglobulins, 3) to devise methods for specific proteolytic cleavage of the myeloma proteins to produce individual domains or functional units such as FV (the combining site composed of VH and VL), and Fc (the locus of biological effector functions), 4) to establish correlations between primary structure conformation, domain structure, and biological effector properties such as complement fixation, and 5) to relate the structure of the monoclonal proteins from individual patients to idiopathic differences in physical and biological properties. New directions include preparation by limited proteolytic cleavage and HPLC of structurally distinct fragments, such as Fc of IgD, and study of their conformation and potential biological activities. Methods to be used include amino acid sequence analysis, structural study of carbohydrates, computer calculations and molecular modelling, physical methods such as CD and NMR, chemical methods such as specific site labeling and carbohydrate structural study, and collaboration with X-ray crystallographers. Another approach is application of a new method for automated tandem HPLC to peptide profile mapping of myeloma proteins, and to preparation of peptides and glycopeptides. This is being applied to IgD, to heavy-chain disease (HCD) proteins of several isotypes, and to allotypic myeloma IgG proteins. One objective is to identify the multiple aberrant proteins made by patients with heavy chain disease and to relate the structure of these proteins to aberrations in gene rearrangements and in protein and carbohydrate processing. Other objectives include structural characterization of carbohydrate variants of myeloma proteins and identification of the sites of allotypic substitutions. Concurrent with these objectives is the goal of facilitating the development of more specific immunochemical reagents for the classification of IgM, IgG, IgD, IgA, and HCD proteins in normal and pathological sera, for detection in tissues, and as cell surface markers.
