Long-range cooperative binding effects in a T cell receptor variable domain

Abstract

Although cellular processes depend on protein-protein interactions, our understanding of molecular recognition between proteins remains far from comprehensive. Protein-protein interfaces are structural and energetic mosaics in which a subset of interfacial residues, called hot spots, contributes disproportionately to the affinity of the complex. These hot-spot residues can be further clustered into hot regions. It has been proposed that binding energetics between residues within a hot region are cooperative, whereas those between hot regions are strictly additive. If this idea held true for all protein-protein interactions, then energetically significant long-range conformational effects would be unlikely to occur. In the present study, we show cooperative binding energetics between distinct hot regions that are separated by >20 Å. Using combinatorial mutagenesis and surface plasmon resonance binding analysis to dissect additivity and cooperativity in a complex formed between a variable domain of a T cell receptor and a bacterial superantigen, we find that combinations of mutations from each of two hot regions exhibited significant cooperative energetics. Their connecting sequence is composed primarily of a single ␤-strand of the T cell receptor variable Ig domain, which has been observed to undergo a strand-switching event and does not form an integral part of the stabilizing core of this Ig domain. We propose that these cooperative effects are propagated through a dynamic structural network. Cooperativity between hot regions has significant implications for the prediction and inhibition of protein-protein interactions.binding energy ͉ cooperativity ͉ protein-protein interaction ͉ surface plasmon resonance ͉ T cell activity

Molecular Architecture of the Major Histocompatibility Complex Class I-binding Site of Ly49 Natural Killer Cell Receptors

“…Such communication between distant sites in proteins has been observed in many systems (33,34), and it appears to involve long range perturbations in electrostatic fields or vibrational modes that are still poorly understood.…”

TCR recognition of peptide/MHC class II complexes and superantigens

“…No contacts with residues from the CDR1, CDR3 or HV4 loops are made with TSST-1. Hot spot residues located from the CDR2 and FR3 on opposite sides of the binding interface act synergistically to bind TSST-1 in an energetically cooperative manner [38].…”

“…Binding to the TCR Vβ CDR2 loop is a requirement for all bacterial SAGs, and the proportion of the SAG-TCR interface that is contributed by the CDR2 loop is invariably the greatest in any SAG-TCR complex, relative to any other single hypervariable or framework region. Involvement of Vβ domain regions beyond the CDR2 loop, however, plays a significant role in the TCR Vβ domain specificity and cross-reactivity of a SAG [38,49,54]. SEK and TSST-1 engage one or more framework region apical loops, at the expense of contacting the hypervariable elements.…”

Energetic determinants of protein binding specificity: Insights into protein interaction networks

“…Alanine scanning analysis has revealed that some binding hot spots are conserved within protein families [19], whereas others are specific to each family member [20][21][22]. Experiments have emphasized the modular design of binding sites, with energetic cooperative contribution of single residues within the module; and additive between modules [23][24][25]. Computational protein design methods have also been used to elucidate the tradeoff between stability and specificity for the optimization of biological function [8,26].…”