Stacking and energetic contribution of aromatic islands at the binding interface of antibody proteins

Abstract

Background: The enrichment and importance of some aromatic residues, such as Tyr and Trp, have been widely noticed at the binding interfaces of antibodies from many experimental and statistical results, some of which were even identified as "hot spots" contributing significantly greater to the binding affinity than other amino acids. However, how these aromatic residues influence the immune binding still deserves further investigation. A large-scale examination was done regarding the local spatial environment around the interfacial Tyr or Trp residues. Energetic contribution of these Tyr and Trp residues to the binding affinity was then studied regarding 82 representative antibody interfaces covering 509 immune complexes from the PDB database and IMGT/3Dstructure-DB.

Results: The connectivity analysis of interfacial residues showed that Tyr and Trp tended to cluster into the spatial Aromatic Islands (AI) rather than being distributed randomly at the antibody interfaces. Out of 82 antibody-antigen complexes, 72% (59) interfaces were found to contain AI with more than 3 aromatic residues. The statistical test against an empirical distribution indicated that the existence of AI was significant in about 60% representative antibody interfaces. Secondly, the loss of solvent accessible surface area (SASA) for side chains of aromatic residues between actually crowded state and independent state was nicely correlated with the AI size increasing in a linearly positive way which indicated that the aromatic side chains in AI tended to take a compact and ordered stacking conformation at the interfaces. Interestingly, the SASA loss of AI was also correlated roughly with the averaged gap of binding free energy between the theoretical and experimental data for immune complexes.

Conclusions: The results of our study revealed the wide existence and statistical significance of "Aromatic Island" (AI) composed of the spatially clustered Tyr and Trp residues at the antibody interfaces. The regular arrangement and stacking of aromatic side chains in AI could probably produce extra cooperative effects to the binding affinity which was firstly observed through the large-scale data analysis. The finding in this work not only provides insights into the functional role of aromatic residues in the antibody-antigen interaction, but also may facilitate the antibody engineering and potential clinical applications.

Figure 1

Flatten view of a binding interface at an antibody (PDB code 1c08). V_LTyr50, V_LTrp94, V_LTyr96, V_HTyr33, V_HTyr50, V_HTyr53, V_HTyr58, V_HTrp98 (IMGT numbering: V-Kappa Tyr56, V-Kappa Trp114, V-Kappa Tyr116, VH Tyr38, VH Tyr55, VH Tyr58, VH Tyr66, VH Trp107) cluster together to form a spatial “Aromatic Island” (abbreviated to “AI”) at the HyHEL-10 antibody interfaces, which is highlighted with white line (Tyr, colored in dark yellow; Trp, colored in orange). Figure 1 is generated by software JMiV.

Figure 2

Correlation between the size of Aromatic Island and the loss of SASA for Aromatic Island The loss of SASA for AI (Ų) is calculated as the difference between the sum of SASA for every side chain of aromatic residue in fully independent state and the actual SASA for Aromatic Island. The size of Aromatic Island is recorded as the number of aromatic Tyr and Trp residues included in AI

Figure 3

Stacking Conformation for aromatic residues in Aromatic Island The antigen protein is displayed with solvent surface mode. For better view, only the aromatic interfacial Tyr and Trp residues in AI are displayed with stick mode from the antibody. (A) Aromatic Island size of 4 at the binding interface of antibody 9D7 and IL-10 (PDB code 1lk3) (B) Aromatic Island size of 5 at the binding interface of antibody 33H1 and potassium channel molecule (PDB code 1ors) (C) Aromatic Island size of 6 at the binding interface of antibody and cytochrome AA3 (PDB code 1ar1) (D) Aromatic Island size of 7 at the binding interface of antibody YTS 105.18 and T-cell surface glycoprotein CD8 alpha chain (PDB code 2arj) (E) Aromatic Island size of 8 at the binding interface of antibody HyHEL-26 and HEL (PDB code 1ndm) (F) Aromatic Island size of 9 at the binding interface of antibody HyHEL-10 mutant and HEL (PDB code 2eiz)

Figure 4

Correlation between the energetic gap and SASA loss of Aromatic Islands at 30 antibody interfaces. Along the x-axis all of the values in a bin (size 200 Ų) are pulled together as a group and shown in the middle. The gap of binding free energy between theoretical and experimental data is indicated with grey square for every immune complex. In each group, the gap is averaged and indicated with black dot.