Dissecting cooperative and additive binding energetics in the affinity maturation pathway of a protein-protein interface.

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TitleDissecting cooperative and additive binding energetics in the affinity maturation pathway of a protein-protein interface.
Publication TypeJournal Article
Year of Publication2003
AuthorsYang, J, Swaminathan, CP, Huang, Y, Guan, R, Cho, S, Kieke, MC, Kranz, DM, Mariuzza, RA, Sundberg, EJ
JournalJ Biol Chem
Volume278
Issue50
Pagination50412-21
Date Published2003 Dec 12
ISSN0021-9258
KeywordsAlgorithms, Animals, Binding Sites, Crystallography, X-Ray, Enterotoxins, Escherichia coli, Kinetics, Mice, Models, Molecular, Mutation, Peptide Fragments, Protein Binding, Protein Structure, Tertiary, Receptors, Antigen, T-Cell, alpha-beta, Sodium Chloride, Static Electricity, Surface Plasmon Resonance, Thermodynamics, Time Factors
Abstract

When two proteins associate they form a molecular interface that is a structural and energetic mosaic. Within such interfaces, individual amino acid residues contribute distinct binding energies to the complex. In combination, these energies are not necessarily additive, and significant positive or negative cooperative effects often exist. The basis of reliable algorithms to predict the specificities and energies of protein-protein interactions depends critically on a quantitative understanding of this cooperativity. We have used a model protein-protein system defined by an affinity maturation pathway, comprising variants of a T cell receptor Vbeta domain that exhibit an overall affinity range of approximately 1500-fold for binding to the superantigen staphylococcal enterotoxin C3, in order to dissect the cooperative and additive energetic contributions of residues within an interface. This molecular interaction has been well characterized previously both structurally, by x-ray crystallographic analysis, and energetically, by scanning alanine mutagenesis. Through analysis of group and individual maturation and reversion mutations using surface plasmon resonance spectroscopy, we have identified energetically important interfacial residues, determined their cooperative and additive energetic properties, and elucidated the kinetic and thermodynamic bases for molecular evolution in this system. The summation of the binding free energy changes associated with the individual mutations that define this affinity maturation pathway is greater than that of the fully matured variant, even though the affinity gap between the end point variants is relatively large. Two mutations in particular, both located in the complementarity determining region 2 loop of the Vbeta domain, exhibit negative cooperativity.

DOI10.1074/jbc.M306848200
Alternate JournalJ. Biol. Chem.
PubMed ID14514664
Grant ListAI49564 / AI / NIAID NIH HHS / United States
GM52801 / GM / NIGMS NIH HHS / United States