Behavior of soluble human 125I-labeled C3b, the third component of complement, after binding to human cells.
The behavior of 125I-labeled C3b incubated with two C3b receptor-positive cells (human erythrocytes and the B lymphoblastoid Raji line), one C3b receptor-negative cell (T lymphoblastoid CEM line) and solubilized membranes from each cell was analyzed by sucrose density gradient (SDG) and sodium dodecyl sulfate polyacrylamide gel electrophoresis. Whichever whole cell was tested, the unbound 125I-labeled C3b recovered in the cell supernatant was not cleaved. When 125I-labeled C3b was bound to whole cells or incubated with solubilized membranes, three different activities were detected: (a) nonspecific C3b polymerization, induced on the membrane of C3b receptor-positive or C3b receptor-negative cells; (b) specific C3b receptor activity solubilized only from the membrane of the two C3b receptor-positive cells and (c) C3b hydrolytic activity, inhibited by 5 X 10(-4)M phenyl methyl sulfonyl fluoride, only extracted from human erythrocyte membranes and carried by a molecule different from that of C3b receptor. C3b receptor activity solubilized from Raji and human erythrocyte membranes was detected by a 12S peak complex formation on a 10-30% SDG and characterized by an affinity constant of 2 X 10(7) to 4 X 10(7) mol-1. Hydrolysis of labeled C3b (Mr = 175000) by solubilized human erythrocyte membranes led to the formation of a split product of Mr = 35000 consisting of two disulfide-linked polypeptide chains of Mr = 17000. This is the first report of a breakdown of C3b on cell membranes different from the physiological breakdown described in the fluid phase.
D001402:B-Lymphocytes / Q000276:immunology / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D003179:Complement C3b / Q000378:metabolism*; D004910:Erythrocyte Membrane / Q000378:metabolism; D004912:Erythrocytes / Q000276:immunology / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D046911:Macromolecular Substances; D011951:Receptors, Complement; D012995:Solubility; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism
D046911:Macromolecular Substances; D011951:Receptors, Complement; D003179:Complement C3b