ENHANCEMENT AND DESTRUCTION OF ANTIBODY FUNCTION BY SOMATIC MUTATION - UNEQUAL OCCURRENCE IS CONTROLLED BY V-GENE COMBINATORIAL ASSOCIATIONS

We examined the positive and negative effects of somatic mutation on a ntibody function using saturation mutagenesis in vitro to mimic the po tential of the in vivo process to diversify antibodies, Identical muta tions were introduced into the second complementarity determining regi on of two anti-phosphocholine antibodies, T15 and D16, which share the same germline VH gene sequence, T15 predominates in primary responses and does not undergo affinity maturation, D16 is representative of an tibodies that co-dominate in memory responses and do undergo affinity maturation, We previously reported that >50% of T15 mutants had decrea sed antigen binding capacity, To test if this high frequency of bindin g loss was unique to T15 or a consequence of random point mutations ap plicable to other combining sites, we analyzed the same mutations in D 16, We show that D16 suffers a similar loss of function, indicating an equally high potential for B-cell wastage, However, only D16 displaye d the capacity for somatic mutation to improve antigen binding, which should enhance its persistence in memory responses, Mutation of residu es contacting the haptenic group, as determined by molecular modeling, did not improve binding, Instead, productive mutations occurred in re sidues that either contacted carrier protein or were distant from the antigen binding site, possibly increasing binding site flexibility thr ough long-range effects, Targeting such residues for mutation should a id in the rational design of improved antibodies.