1jfq
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ANTIGEN-BINDING FRAGMENT OF THE MURINE ANTI-PHENYLARSONATE ANTIBODY 36-71, "FAB 36-71"
OverviewOverview
Alanine scanning was used to determine the affinity contributions of 10, side chain amino acids (residues at position 50-60 inclusive) of H chain, complementarity-determining region 2 (HCDR2) of the somatically mutated, high-affinity anti-p-azophenylarsonate Ab, 36-71. Each mutated H chain, gene was expressed in the context of mutated (36-71L) and the unmutated, (36-65L) L chains to also assess the contribution of L chain mutations to, affinity. Combined data from fluorescence quenching, direct binding, inhibition, and capture assays indicated that mutating H:Tyr(50) and, H:Tyr(57) to Ala in the 36-71 H chain results in significant loss of, binding with both mutated (36-71L) or unmutated (36-65L) L chain, although, the decrease was more pronounced when unmutated L chain was used. All, other HCDR2 mutations in 36-71 had minimal effect on Ab affinity when, expressed with 36-71 L chain. However, in the context of unmutated L, chain, of H:Gly(54) to Ala resulted in significant loss of binding, while, Abs containing Asn(52) to Ala, Pro(53) to Ala, or Ile(58) to Ala mutation, exhibited 4.3- to 7.1-fold reduced affinities. When alanine scanning was, performed instead on certain HCDR2 residues of the germline-encoded, (unmutated) 36-65 Ab and expressed with unmutated L chain as Fab in, bacteria, these mutants exhibited affinities similar to or slightly higher, than the wild-type 36-65. These findings indicate an important role of, certain HCDR2 side chain residues on Ab affinity and the constraints, imposed by L chain mutations in maintaining Ag binding.
About this StructureAbout this Structure
1JFQ is a Protein complex structure of sequences from Mus musculus. Full crystallographic information is available from OCA.
ReferenceReference
Structural analysis of mutants of high-affinity and low-affinity p-azophenylarsonate-specific antibodies generated by alanine scanning of heavy chain complementarity-determining region 2., Parhami-Seren B, Viswanathan M, Strong RK, Margolies MN, J Immunol. 2001 Nov 1;167(9):5129-35. PMID:11673524
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