Prediction of the interacting surfaces in a trimolecular complex formed between the major dust mite allergen Der p 1, a mouse monoclonal anti-Der p 1 antibody, and its anti-idiotype

Abstract

Background: Two mouse monoclonal antibodies (mAbs) have been described recently; namely, mAb 2C7 (IgG2b kappa), which is directed against the major house dust mite allergen Der p 1, and mAb 2G10 (IgG1 kappa), which is an anti-idiotypic antibody raised against mAb 2C7. The anti-idiotype mAb 2G10 does not block the binding of mAb 2C7 to Der p 1, which means that mAb 2C7 can simultaneously bind to Der p 1 and to mAb 2G10, thereby generating a trimolecular complex consisting of antigen-idiotype-anti-idiotype.

Aims: To sequence and model the V region of the anti-idiotypic antibody mAb 2G10 to enable the prediction of the interacting surfaces in the trimolecular complex consisting of Der p 1-mAb 2C7-mAb 2G10.

Methods: DNA sequencing of mAb 2G10 was carried out and the Swiss Model and Swiss PDB-Viewer programs were used to build a three dimensional model of the trimolecular complex.

Results: Complementarity of shape and charge was revealed when comparing the protrusion of the previously determined Der p 1 epitope (Leu147-Gln160) with the cavity formed by the complementarity determining regions (CDRs) of mAb 2C7. Such complementarity was also observed between the mAb 2C7 epitope predicted to be recognised by mAb 2G10 (residues Lys19 from framework region 1 (FRW1) and Ser74-Gln81 from FRW3) and residues from the CDRs of mAb 2G10 (a negatively charged patch flanked by the residues Asp55H/Glu58H and Glu27L/Glu27cL). As expected, the location of the mAb 2C7 epitope recognised by mAb 2G10 does not appear to interfere with the binding of Der p 1 to mAb 2C7.

Conclusion: Although the results obtained represent only an approximation, they nevertheless provide a rare insight into how an antigen (Der p 1) might bind to its antibody (mAb 2C7) while in complex with an anti-idiotype (mAb 2G10).

Figure 1

A 1% agarose gel showing PCR products and restriction digests of plasmids containing the monoclonal antibody 2G10 cDNA inserts for the light chain(top lanes 2–9) and the heavy chain Fd fragment (bottom lanes 2–9). Lane M contains a 100 bp ladder.

Figure 2

Sequence alignments of the monoclonal antibody 2G10 V_L (top panel) and V_H (bottom panel) regions with homologous V_L (1MF2, 1ACY, and 1GGB34) and V_H (1AE6, 1PLG, and 1FBI36) sequences having structural coordinates available within the protein databank. Complementarity determining regions (CDRs) are indicated by solid lines above the sequences; dots indicate missing amino acids.

Figure 3

(A) The predicted epitope on monoclonal antibody (mAb) 2C7 recognised by mAb 2G10, shown in backbone representation (polar residues are shown in cyano blue and non-polar rersidues in red). (B) Side view of the Fv region of mAb 2C7 showing the location of the predicted epitope on framework region 3 (FRW3) of V_H (dark blue). (C) Differences in shape and polarity (shown in space filling representation) between mAb 2G10 reactive (upper row) and non-reactive (lower row) antibodies. Amino acids that are different from those of mAb 2C7 are highlighted

Figure 4

(A) Front and (B) side views of the Der p 1–mAb 2C7 complex showing shape complementarity between the Der p 1 (top structure) epitope recognised by monoclonal antibody (mAb) 2C7 (Leu147–Gln160) and the Fv region of mAb 2C7 (lower structure). The contacting surfaces between (C) Der p 1 and (D) mAb 2C7 are defined by a quadrangular patch (blue), which in Der p 1 is surrounded by residues Arg151, Arg156, Gln118, and Gln166 and in mAb 2C7 by residues Pro61H, Thr5L, Trp103H, and Lys 75H. The previously determined epitope for mAb 2C7 on Der p 1 is shown in purple.

Figure 5

Complementarity of (A) shape and (B and C) electrostatic potential between the monoclonal antibody (mAb) 2C7 epitope on Der p 1 and the Fv region of mAb 2C7. The protrusion formed by the Der p 1 epitope (Leu147–Gln160) fits into the cavity defined by the solvent accessible residues (Lys64H, Arg24L, and Arg97H) of the complementarity determining regions (CDRs) of mAb 2C7. Complementarity of electrostatic potential between mAb 2C7 (a negatively charged patch defined by residues Asp53H, Asp53L, and Asp28H) (B) and Der p 1 (a positively charged patch defined by residues Arg151, Arg156, Arg161, and Lys145) (C) is shown in red and blue, respectively. The mAb 2C7 residues Asp53H and Asp28H (B) are complementary to the Der p 1 residues Arg156 and Arg161 (C), respectively. The protruding Der p 1 residues Lys145 and Arg151 fit into two cavities formed between the mAb 2C7 residues Asp28H and Asp53H on one side and Asp28H, Asp53H, and Asp53L on the other side. The Fv region of mAb 2C7 (A) is coloured according to solvent accessibility, decreasing in the order yellow, green, light blue, and dark blue. The electrostatic potential was computed by the program Swiss PDB-Viewer, which uses simple coulomb interaction.

Figure 6

A molecular model of the trimolecular complex consisting of Der p1–monoclonal antibody (mAb) 2C7–mAb 2G10. The previously determined Der p 1 epitope recognised by mAb 2C7 (Leu147–Gln160), shown in space filling representation (purple). The mAb 2C7 epitope predicted to be recognised by mAb 2G10 forms a protrusion, involving residues Ser74–Gln81 (framework region 3 (FRW3)) and Lys19 (FRW1), which fits into a cavity, defined by residues Asp55H and Glu58H, on the complementarity determining regions (CDRs) of mAb 2G10. (A) Side and (B) front views of the predicted mAb 2G10 epitope on mAb 2C7 and the CDRs of mAb 2G10, shown in space filling representation and coloured according to solvent accessibility, decreasing in the order yellow, green, light blue, and dark blue. (C) Side and (D) front views of the trimolecular complex represented by its molecular surfaces and coloured according to electrostatic potential. Again, the electrostatic potential was computed by the program Swiss PDB-Viewer, which uses simple coulomb interaction. The mAb 2C7 epitope predicted to be recognised by mAb 2G10 consists of a positively charged protrusion (blue), which fits into a negatively charged cavity (red) on the CDRs of mAb 2G10. The positively charged mAb 2C7 residues Lys75H and Lys19H seem to fit into two cavities formed between the mAb 2G10 negatively charged residues (coloured pink) Asp55H/Glu58H and Glu27L/Glu27cL, respectively. In (A) and (C) mAb 2G10 is pulled apart from the Der p 1–mAb 2C7 complex and viewed from the side. In (B) and (D) each component is rotated 90° towards the viewer.