A complement- (C) fixing particle consisting of agarose beads to which 5-thioglucose was attached by a -S-S-bond (agarose-thioglucose) was employed to investigate the mechanism of attachment of C3 to surfaces. When whole serum containing [ 125I] C3 was incubated with agarose-thioglucose, labeled C3b was taken up in a form that was not removed by 2 M NaCl but was released by 10 mM dithiothreitol. Deposition of DDT-releasable C3b was dependent upon the alternative pathway of C activation. Gel electrophoresis of DTT-releasable C3b from similar experiments was dependent upon the alternative pathway of C activation. Gel electrophoresis of DTT-releasable C3b from similar experiments performed with unlabeled serum and agarose-[ 3H]thioglucose showed that the liberated C3b contained a molecule of radioactive thioglucose attached to the α'-chain by a covalent bond that was stable to mercaptoethanol. We propose that the thioglucose-α' chain bond was formed during the course of C activation by a reaction between the 'labile binding site' of newly released C3b and the (then) particle-bound sugar. This formulation implies that the reaction by which C3b attaches to 5-thioglucose in this system is the reaction responsible for opsonization by C3b, and that the C3b-linked sugar represents a marker for the labile binding site. Incubation of the particle-bound C3b in serum resulted in the cleavage of the covalently linked α'-chain to several smaller polypeptides, the major cleavage product having a m.w. of 70,000.
|Original language||English (US)|
|Number of pages||3|
|Journal||Journal of Immunology|
|State||Published - 1981|
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