Analysis of the mechanism, rate, and sites of proteolytic cleavage of human immunoglobulin D by high-pressure liquid chromatography.
The high susceptibility of human immunoglobulin D to proteolytic degradation affects its biological function, metabolism, and immunoassay. High-pressure liquid chromatography was used to investigate the mechanism and rate of limited proteolytic cleavage of IgD and also to identify, isolate, and quantify the reaction products. Within 1 to 5 min, tryptic digestion of native IgD almost quantitatively yields a labile Fab fragment, a stable Fc fragment, and a highly charged peptide derived from the hinge region. A galactosamine-rich glycopeptide from the hinge region increases inversely as the Fab is largely degraded to a series of peptides within 1 hr. In contrast, the Fc and the high-charge peptide resist proteolysis for more than 24 hr. The initial sites of cleavage of IgD occur in the hinge region at exposed secondary structures predicted to be beta-turns. Concomitant with removal of the galactosamine-rich glycopeptide at its carboxyl terminus, the Fd fragment is rapidly and rather randomly degraded, but the light chain is somewhat more resistant than the Fd section of the delta heavy chain. This study of the rapid rate of proteolysis of IgD explains the rarity with which intact IgD is found in human sera. It also raises questions about immunoassay of IgD, which is usually measured with antisera against Fc. In vivo, proteolytic cleavage initiates the catabolism of circulating IgD and also affects the role and fate of IgD as an antigen receptor on the B-cell membrane.
Ishioka
N
N
Takahashi
N
N
Putnam
F W
FW
D002851:Chromatography, High Pressure Liquid; D006020:Glycopeptides / Q000032:analysis; D006801:Humans; D007072:Immunoglobulin D / Q000378:metabolism*; D007140:Immunoglobulin Fab Fragments / Q000032:analysis; D007141:Immunoglobulin Fc Fragments / Q000032:analysis; D007128:Immunoglobulin Fragments / Q000032:analysis; D007700:Kinetics; D014357:Trypsin / Q000378:metabolism
D006020:Glycopeptides; D007072:Immunoglobulin D; D007140:Immunoglobulin Fab Fragments; D007141:Immunoglobulin Fc Fragments; D007128:Immunoglobulin Fragments; D014357:Trypsin