Molecular basis for the temperature-dependent insolubility of cryoglobulins-IV. Structural studies of the IgM monoclonal cryoglobulin McE

A 20.1s⁰_2,w monoclonal IgM, κ immunoglobulin (McE) exhibiting reversible, temperaturedependent insolubility is described. The precipitation of the protein upon reduction in solution temperature displays sharp concentration dependence and is optimal between pH 6 and 8. The effect of temperature on the rate of cryoprecipitation suggests the involvement of either a low temperature-induced conformational change or nucleation event in cryoprecipitation. Structural studies have indicated that the masses (under denaturing conditions) of both heavy and light chains derived from the cryoimmunoglobulin and noncryoimmunoglobulin reference proteins are identical, and the protein possesses a characteristic J chain. The cryoimmunoglobulin is converted to (Fc)₅μ§ and Fabμ fragments by limited digestion with trypsin at 56° C. The heavy chain of the McE protein is deficient in tyrosine and contains excess tryptophan relative to noncryoglobulin and other cryoglobulin proteins. Calculation of structural parameters from these amino acid compositions suggests that the heavy chains of cryoimmunoglobulins may contain significantly greater (P< 0.01) amounts of polypeptide chain turns than do noncryoglobulin references. With the exception of a Leu → Phe substitution at position 13, the McE light chain is homologous with other immunoglobulins of the V_κII subgroup, while the heavy chain is of the less frequently encountered V_HV subgroup. (Fc)₅μ and Fabμ fragments, as well as reduction-derived monomeric subunits, do not cryoprecipitate. Removal of J chain does not affect cryoprecipitation of the reassembled molecule while partial enzymatic removal of neutral hexoses and glucosamine enhances cryoprecipitation. Cryoprecipitation of McE is interpreted as a general solubility effect as opposed to an antibody-antigen phenomenon.