PDBsum entry 1jfq

Immune system

PDB id: 1jfq

Contents

Protein chains

215 a.a. *

220 a.a. *

Waters ×112

* Residue conservation analysis

PDB id:

1jfq

Name:

Immune system

Title:

Antigen-binding fragment of the murine anti-phenylarsonate antibody 36-71, "fab 36-71"

Structure:

Antigen-binding fragment of anti-phenylarsonate antibody. Chain: l. Fragment: light chain. Engineered: yes. Antigen-binding fragment of anti-phenylarsonate antibody. Chain: h. Fragment: heavy chain. Engineered: yes

Source:

Mus musculus. House mouse. Organism_taxid: 10090. Expressed in: mus musculus. Expression_system_taxid: 10090. Expression_system_cell_line: hybridoma cells. Expression_system_taxid: 10090

Biol. unit:

Dimer (from PQS)

Resolution:

1.90Å R-factor: 0.196

Authors:

B.Parhami-Seren,M.Viswanathan,R.K.Strong,M.N.Margolies

Key ref:

B.Parhami-Seren et al. (2001). 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. J Immunol, 167, 5129-5135. PubMed id: 11673524

Date:

21-Jun-01 Release date: 27-Feb-02

Protein chain

P01648  (KV5AF_MOUSE) -  Ig kappa chain V-V region HP 91A3 from Mus musculus

Seq: 108 a.a.

Struc: 215 a.a.*

Protein chain

P01747  (HVM03_MOUSE) -  Ig heavy chain V region 36-65 from Mus musculus

Seq: 120 a.a.

Struc: 220 a.a.*

* PDB and UniProt seqs differ at 17 residue positions (black crosses)

J Immunol 167:5129-5135 (2001)

PubMed id: 11673524

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.

B.Parhami-Seren, M.Viswanathan, R.K.Strong, M.N.Margolies.

ABSTRACT

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.