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

BackgroundThe 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.ResultsThe 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.ConclusionsThe 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.

[1]  Chantal Abergel,et al.  Identification of specificity‐determining residues in antibodies , 1995, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[2]  D. Baker,et al.  Design of a Novel Globular Protein Fold with Atomic-Level Accuracy , 2003, Science.

[3]  Mathieu Rouard,et al.  IMGT unique numbering for immunoglobulin and T cell receptor constant domains and Ig superfamily C-like domains. , 2005, Developmental and comparative immunology.

[4]  Hongyi Zhou,et al.  A physical reference state unifies the structure‐derived potential of mean force for protein folding and binding , 2004, Proteins.

[5]  V. Giudicelli,et al.  IMGT unique numbering for immunoglobulin and T cell receptor variable domains and Ig superfamily V-like domains. , 2003, Developmental and comparative immunology.

[6]  Shohei Koide,et al.  The importance of being tyrosine: lessons in molecular recognition from minimalist synthetic binding proteins. , 2009, ACS chemical biology.

[7]  L. Jin,et al.  High resolution functional analysis of antibody-antigen interactions. , 1992, Journal of molecular biology.

[8]  S. Jones,et al.  Analysis of protein-protein interaction sites using surface patches. , 1997, Journal of molecular biology.

[9]  E. Padlan,et al.  Anatomy of the antibody molecule. , 1994, Molecular immunology.

[10]  Marie-Paule Lefranc,et al.  IMGT/3Dstructure-DB and IMGT/StructuralQuery, a database and a tool for immunoglobulin, T cell receptor and MHC structural data , 2004, Nucleic Acids Res..

[11]  C. Chothia,et al.  The atomic structure of protein-protein recognition sites. , 1999, Journal of molecular biology.

[12]  Yili Li,et al.  Dissection of binding interactions in the complex between the anti-lysozyme antibody HyHEL-63 and its antigen. , 2003 .

[13]  D. Stuart,et al.  Analysis of antigenic surfaces of proteins , 1995, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[14]  Marie-Paule Lefranc,et al.  IMGT/3Dstructure-DB and IMGT/DomainGapAlign: a database and a tool for immunoglobulins or antibodies, T cell receptors, MHC, IgSF and MhcSF , 2009, Nucleic Acids Res..

[15]  Peter A. Kollman,et al.  Calculating Structures and Free Energies of Complex Molecules: Combining Molecular Mechanics and Continuum Models , 2001 .

[16]  N. Ben-Tal,et al.  Residue frequencies and pairing preferences at protein–protein interfaces , 2001, Proteins.

[17]  T. Bhat,et al.  Solvent rearrangement in an antigen-antibody interface introduced by site-directed mutagenesis of the antibody combining site. , 1994, Journal of molecular biology.

[18]  B. Lee,et al.  The interpretation of protein structures: estimation of static accessibility. , 1971, Journal of molecular biology.

[19]  W. Saenger,et al.  Residue solvent accessibilities in the unfolded polypeptide chain. , 1992, Biophysical journal.

[20]  G. M. Griffiths,et al.  Molecular events during maturation of the immune response to oxazolone , 1985, Nature.

[21]  H. Bilofsky,et al.  Unusual distributions of amino acids in complementarity-determining (hypervariable) segments of heavy and light chains of immunoglobulins and their possible roles in specificity of antibody-combining sites. , 1977, The Journal of biological chemistry.

[22]  Richard H. Henchman,et al.  Revisiting free energy calculations: a theoretical connection to MM/PBSA and direct calculation of the association free energy. , 2004, Biophysical journal.

[23]  A. Bogan,et al.  Anatomy of hot spots in protein interfaces. , 1998, Journal of molecular biology.

[24]  M. Gilson,et al.  The statistical-thermodynamic basis for computation of binding affinities: a critical review. , 1997, Biophysical journal.

[25]  A. D. de Vos,et al.  Selection and analysis of an optimized anti-VEGF antibody: crystal structure of an affinity-matured Fab in complex with antigen. , 1999, Journal of molecular biology.

[26]  Sachdev S Sidhu,et al.  Tyrosine plays a dominant functional role in the paratope of a synthetic antibody derived from a four amino acid code. , 2006, Journal of molecular biology.

[27]  W. Delano.  Unraveling hot spots in binding interfaces: progress and challenges. , 2002, Current opinion in structural biology.

[28]  Nikolai Petrovsky,et al.  Immunome Research , 2005, Immunome research.

[29]  K. Tsumoto,et al.  Structural Consequences of Mutations in Interfacial Tyr Residues of a Protein Antigen-Antibody Complex , 2007, Journal of Biological Chemistry.

[30]  S. Smith‐Gill,et al.  Dissection of binding interactions in the complex between the anti-lysozyme antibody HyHEL-63 and its antigen. , 2003, Biochemistry.

[31]  S. Wetmore,et al.  Characterization of the stacking interactions between DNA or RNA nucleobases and the aromatic amino acids , 2007 .

[32]  J. SantaLucia,et al.  The thermodynamics of DNA structural motifs. , 2004, Annual review of biophysics and biomolecular structure.

[33]  M. Gilson,et al.  Calculation of protein-ligand binding affinities. , 2007, Annual review of biophysics and biomolecular structure.

[34]  Sarah A. Teichmann,et al.  Principles of protein-protein interactions , 2002, ECCB.

[35]  Sachdev S Sidhu,et al.  Synthetic antibodies from a four-amino-acid code: a dominant role for tyrosine in antigen recognition. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[36]  Kurt S. Thorn,et al.  ASEdb: a database of alanine mutations and their effects on the free energy of binding in protein interactions , 2001, Bioinform..

[37]  Doheon Lee,et al.  Specificity of molecular interactions in transient protein–protein interaction interfaces , 2006, Proteins.

[38]  A. D. de Vos,et al.  VEGF and the Fab fragment of a humanized neutralizing antibody: crystal structure of the complex at 2.4 A resolution and mutational analysis of the interface. , 1998, Structure.

[39]  I. Mian,et al.  Structure, function and properties of antibody binding sites. , 1991, Journal of molecular biology.

[40]  T. Manser,et al.  Somatic evolution of variable region structures during an immune response. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[41]  B. Braden,et al.  Estimation of the hydrophobic effect in an antigen-antibody protein-protein interface. , 2000, Biochemistry.

[42]  P. Kollman,et al.  Calculating structures and free energies of complex molecules: combining molecular mechanics and continuum models. , 2000, Accounts of chemical research.

[43]  J. C. Cheetham,et al.  Generation of an antibody with enhanced affinity and specificity for its antigen by protein engineering , 1987, Nature.

[44]  Peter A. Kollman,et al.  FREE ENERGY CALCULATIONS : APPLICATIONS TO CHEMICAL AND BIOCHEMICAL PHENOMENA , 1993 .

[45]  G. Cohen,et al.  Interactions of protein antigens with antibodies. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[46]  T. N. Bhat,et al.  The Protein Data Bank , 2000, Nucleic Acids Res..