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Having investigated relationships between antibody sequences, antigen contacts, antigen type and combining site topography, we applied these results by predicting the antigen-binding pockets of antibody structures. This is particularly relevant for antibodies whose sequences are known and from which a model can be constructed. From the model we aim to predict those residues most likely to be involved in antigen contact, which can then be subjected to site-directed mutagenesis to test both theory and model.
We have developed a rapid and simple method which uses only average contact and surface shape information and test it by application to the complexed crystal structures (all antigen-contact data from the antibody being predicted are excluded). Firstly the mean burial (by antigen) data, , for each residue is projected onto the molecular surface of the antibody (Figure 2.7(a)). Figure 2.7(b) shows the same antibody with the surface coloured according to its curvature, , as calculated by the program GRASP[Nicholls et al., 1991]. This alternative description of curvature was adopted for purely practical reasons and allows easy application of the technique by anyone with access to GRASP. To find the probable contact surface we calculate , which combines the measure of concavity with the probability of antigen contact. An arbitrary cutoff of was chosen by visual inspection (using just one complex), and surface points satisfying this condition are coloured red (Figure 2.7(c)). This red surface is patchy and discontinuous, so neighbouring patches are merged and disconnected patches are eliminated using a dilation/erosion procedure[Delaney, 1992] developed within GRASP. The red surfaces are dilated five times, eroded eight times and dilated again three times (each by 1Å). The binding pocket prediction is the resultant smooth red patch (Figure 2.7(d)).
The `worst case' accuracy, , and `fraction correct' accuracy, ,
for predicting antigen contacting residues by this protocol with each of
the 26 complexed crystal structures are calculated as follows: