DELETION IN HCDR3 RESCUES T15 ANTIBODY MUTANTS FROM A SECRETION DEFECT CAUSED BY MUTATIONS IN HCDR2

We recently described mutants oi the murine anti-phosphocholine Ab T15 , with changes in heavy chain complementarity determining region 2 (HC DR2) that caused loss of secretion, Surprisingly, the T15 HCDR2 mutati ons did not alter secretion when placed into the related anti-phosphoc holine Ab D16, which differs from T15 only in HCDR3 and light (L) chai n, Here, we exploit the differences between these two Abs to assess th e basis of the secretion defect, The T15 L chain is not secreted in th e absence of heavy (H) chain, In contrast, D16 L chain is secreted in the absence of H chain, as are most L chains, We co-expressed the T15 wild-type (wt) and mutant H chains with the D16 L chain, as well as wi th another secreted L chain, J558L, The mutant H chains were not secre ted when expressed with either heterologous L chain, These results est ablish that the T15 L chain is not uniquely associated with the defect , The T15 and D16 Abs also differ in HCDR3 length in that D16 lacks fo ur amino acid residues (Ser(99), Ser(100), Tyr(10Ca), Trp(100b)) prese nt in T15, We deleted these four residues from T15 wt and mutant H cha ins, Secretion of T15 wt was unaffected by the deletion, but shortenin g HCDR3 restored secretion in the HCDR2 mutants regardless of L chain association, Together these data demonstrate that both the HCDR2 and H CDR3 domains contain structural information that may affect the secret ion competence of Abs.