Crystal structure and thermodynamic analysis of diagnostic mAb 106.3 complexed with BNP 5‐13 (C10A) |
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Authors: | Kenton L Longenecker Qiaoqiao Ruan Elizabeth H Fry Sylvia C Saldana Susan E Brophy Paul L Richardson Sergey Y Tetin |
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Institution: | 1. Advanced Technology, Global Pharmaceutical Research and Development, Abbott, Abbott Park, Illinois 60064;2. Diagnostics Research, Diagnostics Division, Abbott, Abbott Park, Illinois 60064 |
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Abstract: | B‐type natriuretic peptide (BNP) is a naturally secreted regulatory hormone that influences blood pressure and vascular water retention in human physiology. The plasma BNP concentration is a clinically recognized biomarker for various cardiovascular diseases. Quantitative detection of BNP can be achieved in immunoassays using the high‐affinity monoclonal IgG1 antibody 106.3, which binds an epitope spanning residues 5‐13 of the mature bioactive peptide. To understand the structural basis of this molecular recognition, we crystallized the Fab fragment complexed with the peptide epitope and determined the three‐dimensional structure by X‐ray diffraction to 2.1 Å resolution. The structure reveals the detailed interactions that five of the complementarity‐determining regions make with the partially folded peptide. Thermodynamic measurements using fluorescence spectroscopy suggest that the interaction is enthalpy driven, with an overall change in free energy of binding, ΔG = ?54 kJ/mol, at room temperature. The parameters are interpreted on the basis of the structural information. The kinetics of binding suggest a diffusion‐limited mechanism, whereby the peptide easily adopts a bound conformation upon interaction with the antibody. Moreover, comparative analysis with alanine‐scanning results of the epitope explains the basis of selectivity for BNP over other related natriuretic peptides. Proteins 2009. © 2009 Wiley‐Liss, Inc. |
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Keywords: | brain natriuretic peptide (BNP) diagnostic biomarker antibody X‐ray structure antibody‐antigen interactions thermodynamic analysis diffusion‐limited mechanism |
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