Isolation and Characterization of the Immune Macroglobulin from the Paddlefish, Polyodon spathula

• Published in: The Journal of biological chemistry 1971-11, Vol.246 (22), p.6760-6769
• Main Authors: Acton, R T, Weinheimer, P F, Dupree, H K, Russell, T R, Wolcott, M, Evans, E E, Schrohenloher, R E, Bennett, J C
• Format: Article
• Language: English
• Subjects: Agglutination Tests; Alkylation; Amino Acid Sequence; Amino Acids - analysis; Animals; Antigens; Biological Evolution; Chromatography, Gel; Chromatography, Ion Exchange; Fishes - immunology; Fucose - analysis; Galactose - analysis; Glucosamine - analysis; Guanidines; Immune Sera; Immunodiffusion; Immunoelectrophoresis; Immunoglobulins; Macroglobulins - analysis; Macroglobulins - isolation & purification; Mannose - analysis; Methods; Molecular Weight; Neuraminic Acids - analysis; Oxidation-Reduction; Peptides - analysis; Protein Conformation; Rabbits; Salmonella typhi; Species Specificity; Spectrophotometry; Trypsin; Ultracentrifugation; Ultraviolet Rays
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(19)45910-3

The immune macroglobulin from the paddlefish, Polyodon spathula , was purified and characterized physicochemically. The macroglobulin had a sedimentation coefficient of 14.2 S and a molecular weight of 660,000. Total reduction and alkylation in 7 m guanidine hydrochloride resulted in dissociation into heavy (H) and light (L) chains. The H and L chains were separated by gel filtration and found to have molecular weights of 58,100 and 21,000, respectively, employing sedimentation equilibrium in 5.0 m guanidine hydrochloride. Based on absorbance, equimolar quantities of the chains were recovered from the macroglobulin. These data are compatible with previously reported electron microscopic studies (A cton , R. T., W einheimer , P. F., D upree , H. K., R ussell , T. R., W olcott , M., E vans , E. E., S chrohenloher , R. E., and B ennett , J. C., Proc. Nat. Acad. Sci. U. S. A. , 68, 107 (1971)), which demonstrated a tetrameric structure for the immune macroglobulin, and indicated that the structural subunit consists of two H and two L chains. Generally the amino acid composition was similar to mammalian IgM H and L chains. Although no NH 2 -terminal group was detected in the light chain, the sequence of the first 5 residues from the NH 2 -terminus of the H chain was found to be [see PDF for sequence]. These data provide additional evidence for a structural similarity among phylogenetically distinct immune macroglobulins and suggest a common evolutionary origin.