A genetically engineered human IgG with limited flexibility fully initiates cytolysis via complement.
A causal link between antigen induced conformational change and functional activity has been previously suggested for the triggering of immunoglobulin effector functions. To test this hypothesis an immunoglobulin with greatly restricted Fab flexibility has been created by site-directed mutagenesis. Serine119, a residue at the elbow in the first heavy chain constant domain of a mouse-human chimeric immunoglobulin, was replaced by cysteine119. The resulting mutant immunoglobulin had an additional intramolecular disulfide bond connecting the two heavy chains. This newly introduced covalent bond between antigen binding arms gave rise to a compact 'tethered' conformation which displayed lowered segmental flexibility as determined by nanosecond fluorescence depolarization anisotropy. However, the ability of this tethered immunoglobulin to initiate lysis of target cells via the complement cascade was unimpaired. Therefore, it is likely that allosteric or distortive mechanisms of conformational change induced complement activation which require unhindered 'twist' and/or 'waggle' motions of Fab are untenable.
Shopes
B
B
Department of Cell Biology, Stanford School of Medicine, CA 94305.
D000818:Animals; D000920:Antibody-Dependent Cell Cytotoxicity; D001483:Base Sequence; D002460:Cell Line; D002678:Chimera; D002850:Chromatography, Gel; D003167:Complement Activation / Q000276:immunology*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000276:immunology; D007074:Immunoglobulin G / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry; D007147:Immunoglobulin Light Chains / Q000737:chemistry; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D013997:Time Factors; D014162:Transfection
D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains