Evidence showing that the 1105 and 1106 isotypic residues of the fourth component of human complement, C4A, are not involved in amide bond formation.
Human C4A and C4B have different functions that may stem from their ability to bind hydroxyl or free amino groups on complement activating surfaces. Previous studies suggest that C4B binds to hydroxyl or amino groups whereas C4A binds to free amino groups on acceptor molecules. Comparison of the derived amino acid sequences of C4A and C4B has shown that differences exist between them at positions 1101, 1102, 1105 and 1106. These residues appear to be involved in the binding specificity of C4B. Less is known about the corresponding residues of C4A. It has been suggested that the aspartic acid of C4A at position 1106 is involved in amide bond formation by serving as a catalytic residue for the reaction or by promoting an increased interaction with amino nucleophilic groups. To examine the functional role of residues 1101-1106, we studied the effects of the C4A site-specific antipeptide mAb, AII-1 in assays dependent on the covalent binding properties of C4A; the C4 mediated inhibition of hemolysis and the C4 mediated inhibition of immune precipitation. This study shows that mAb AII-1 has no effect on C4-mediated hemolysis or its ability to inhibit the rate of immune precipitate formation. The lack of interference by AII-1 in these assays could not be explained by low affinity interaction between antibody and C4A showing that mAb AII-1 does not affect the covalent binding activity of C4A. Furthermore, results from epitope mapping studies show that AII-1 binds to Leu1105 and Asp1106 suggesting that these residues are not critical for amide bond formation by C4A.
Reilly
B D
BD
Department of Microbiology, University of New Mexico, School of Medicine, Albuquerque 87131.
Skanes
V M
VM
Levine
R P
RP
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex / Q000378:metabolism; D001666:Binding Sites, Antibody; D015934:Complement C4a / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes; D005434:Flow Cytometry; D006168:Guinea Pigs; D006461:Hemolysis / Q000276:immunology*; D006801:Humans; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D011485:Protein Binding
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D000939:Epitopes; D015934:Complement C4a