FUNCTIONAL AND MODELING STUDIES OF THE BINDING OF HUMAN MONOCLONAL ANTI-DNA ANTIBODIES TO DNA

The relationships between the antigen-binding specificities of four hu man monoclonal anti-DNA antibodies and the structural aspects of the c ombining sites of two of these were examined. Competition ELISAs were used to examine the reactivities of two IgM MAbs (WRI-176 and RT-79) a nd two IgG mAbs (D5 and B3) to a wide range of polynucleotides. The mA bs WRI-176 and RT-79 were found to bind predominantly ssDNA, with a pr eference for poly (dT), whilst D5 and B3 bound components of both ss- and dsDNA, and Z-DNA. The mAb B3 also exhibited a preference for AT ri ch nucleotides. Computer models were generated for the Fv regions of W RI-176 and B3. Models for RT-79 and D5 were not generated as the struc ture of the long CDR-H3 loops in these mAbs could not be predicted. Th e B3 combining site contains a groove flanked by three arginines at po sitions CDR-L1-27A, CDR-L2-54 and CDR-H2-53. Using interactive molecul ar graphics, B-DNA was docked into the B3 antigen combining site along the plane of the V-H/V-L interface, whilst Z-DNA was best-fitted at a pproximately 90 degrees to this direction. The models provide a hypoth esis to explain the ability of a single autoantibody to bind two diffe rent antigens. In addition, aspects of the base specificity of B3 may be explained. The model of the WRI-176 Fv region revealed a relatively flat surface, on which a large number of hydrophobic and aromatic res idues were present. Trp-H52, in particular, is prominent on the surfac e. This may participate in ssDNA binding through base stacking interac tions. The models allow identification of potential targets for site-d irected mutagenesis. Copyright (C) 1996 Elsevier Science Ltd.