Structure, dynamics, and binding thermodynamics of the v‐Src SH2 domain: Implications for drug design

Taylor, J.G.; Ababou, Abdessamad; Fawaz, Radwan R.; Hobbs, Christopher; Williams, Mark A.; Ladbury, John E.

doi:10.1002/prot.22119

Cited by 22 publications

(39 citation statements)

References 62 publications

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“…This is consistent with the observation that phosphopeptides hardly bind to the ZAP-70 cSH2, whereas phosphate does so readily, in contrast to the cSH2 domain of Syk and to the nSH2 domains of both proteins (6,8,20,21). Structural flexibility is thought to relate to ligand diversity in other SH2 domains (22). For example, a similar mechanism of SH2 gating was recently suggested to regulate SHIP2 (23), differentiating it functionally from the closely related phosphatase SHIP1.…”

Section: Discussionsupporting

confidence: 85%

Molecular Mechanism of Selective Recruitment of Syk Kinases by the Membrane Antigen-Receptor Complex

Bond¹,

Faraldo‐Gómez

2011

Journal of Biological Chemistry

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ZAP-70 and Syk are essential tyrosine kinases in intracellular immunological signaling. Both contain an inhibitory SH2 domain tandem, which assembles onto the catalytic domain. Upon binding to doubly phosphorylated ITAM motifs on activated antigen receptors, the arrangement of the SH2 domains changes. From available structures, this event is not obviously conducive to dissociation of the autoinhibited complex, yet it ultimately translates into kinase activation through a mechanism not yet understood. We present a comprehensive theoretical study of this molecular mechanism, using atomic resolution simulations and free-energy calculations, totaling >10 s of simulation time. Through these, we dissect the microscopic mechanism coupling stepwise ITAM engagement and SH2 tandem structural change and reveal key differences between ZAP-70 and Syk. Importantly, we show that a subtle conformational bias in the inter-SH2 connector causes ITAM to bind preferentially to kinase-dissociated tandems. We thus propose that phosphorylated antigen receptors selectively recruit kinases that are uninhibited and that the resulting population shift in the membrane vicinity sustains signal transduction.Non-receptor Syk tyrosine kinases are an important class among eukaryotic signaling enzymes (1). The family consists of ZAP-70 and Syk, which are mostly found in T-and B-cells, respectively. These kinases help to propagate antigen-recognition signals initiated at the cell membrane. To do so, they first associate with the antigen-receptor complex and then phosphorylate downstream signaling and scaffolding proteins (2). Syk kinases are autoinhibited when isolated; activation follows their engagement of so-called ITAM motifs on the intracellular side of the antigen-receptor complex, through a molecular mechanism that is only partially understood. Consistent with their important cellular role, malfunction of Syk kinases underlies a range of immune system disorders (2, 3).Both ZAP-70 and Syk contain a tandem of SH2 modules, which inhibit the catalytic activity of the enzyme by assembling onto the kinase domain. The SH2 tandem (tSH2) 3 also serves as the docking platform for ITAM motifs. These motifs consist of two repeats of sequence YXX(L/I) connected by a spacer of variable length; doubly phosphorylated ITAM motifs are thus suitable targets for SH2 tandems. Moreover, this dual interaction results in greater affinity, typically in the nanomolar range; that of individual SH2 domains is 2 orders of magnitude weaker (4).Despite the high sequence identity of Syk and ZAP-70 (ϳ60%), they differ noticeably in the thermodynamics of ITAM recognition (5-8). From a functional standpoint, this differentiation is likely related to the fact that Syk is more widely expressed, including in non-immune cells; the diversity of its ligands is therefore necessarily broader (3). However, in view of the similarity of the available tSH2 structures (9, 10), the molecular basis for this differentiation is not immediately clear.The molecular mechanism by which ITAM reco...

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Section: Discussionsupporting

confidence: 85%

Molecular Mechanism of Selective Recruitment of Syk Kinases by the Membrane Antigen-Receptor Complex

Bond¹,

Faraldo‐Gómez

2011

Journal of Biological Chemistry

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“…Nonetheless, the internal dynamics of the pYEEI, cpYEEI, and fpYEEI Src SH2 complexes estimated from 15 N relaxation (Figure 2) and MD fluctuations (Figure 3) show only minor variations, which cannot convincingly account for the observed disparities in binding enthalpy of the three complexes. Previously reported NMR relaxation measurements for Src SH2 also suggest that main-chain flexibility varies little upon binding the canonical pYEEI peptide, with only minimal differences from residues scattered throughout the protein in the amplitude of motion specified from the Lipari-Szabo order parameter54. As such, we find no evidence that alternative conformational distributions occurring from ps-ns motions affect the time-average interaction energy with the three Src SH2 ligands.…”

Section: Discussionsupporting

confidence: 50%

“…The residue is universally conserved across SH2 domains. The structural importance of the hydrogen bonding between R175 and H201 has been supported by the acidic pKa of H201 Nδ, measured by NMR titration in Src SH254 and PLCγ72. A triad of hydrogen bonds between R175, H201, and E159 is argued to preorganize the R175 side-chain in its bound state geometry54.…”

Section: Discussionmentioning

confidence: 97%

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Constraining Binding Hot Spots: NMR and Molecular Dynamics Simulations Provide a Structural Explanation for Enthalpy−Entropy Compensation in SH2−Ligand Binding

et al. 2010

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NMR spectroscopy and molecular dynamics (MD) simulations were used to probe the structure and dynamics of complexes of three phosphotyrosine-derived peptides with the Src SH2 domain in an effort to uncover a structural explanation for enthalpy-entropy compensation observed in the binding thermodynamics. The series of phosphotyrosine peptide derivatives comprises the natural pYEEI Src SH2 ligand, a constrained mimic, in which the phosphotyrosine (pY) residue is preorganized in the bound conformation for the purpose of gaining an entropic advantage to binding, and a flexible analog of the constrained mimic. The expected gain in binding entropy of the constrained mimic was realized; however, a balancing loss in binding enthalpy was also observed that could not be rationalized from the crystallographic structures. We examined protein dynamics to evaluate whether the observed enthalpic penalty might be the result of effects arising from altered motions in the complex. 15N-relaxation studies and positional fluctuations from molecular dynamics indicate that the main-chain dynamics of the protein show little variation among the three complexes. Root mean squared (RMS) coordinate deviations vary by less than 1.5 Å for all non-hydrogen atoms for the crystal structures and in the ensemble average structures calculated from the simulations. In contrast to this striking similarity in the structures and dynamics, there are a number of large chemical shift differences from residues across the binding interface, but particularly from key Src SH2 residues that interact with pY, the ‘hot spot’ residue, which contributes about half of the binding free energy. Rank order correlations between chemical shifts and ligand binding enthalpy for several pY-binding residues, coupled with available mutagenesis and calorimetric data, suggest that subtle structural perturbations (< 1 Å) from the conformational constraint of the pY residue sufficiently alter the geometry of enthalpically critical interactions in the binding pocket to cause the loss of binding enthalpy, leading to the observed entropy-enthalpy compensation. We find no evidence to support the premise that enthalpy-entropy compensation is an inherent property and conclude that preorganization of Src SH2 ligand residues involved in binding hot spots may eventuate in suboptimal interactions with the domain. We propose that introducing constraints elsewhere in the ligand could minimize entropy-enthalpy compensation effects. The results illustrate the utility of the NMR chemical shift to highlight small, but energetically significant, perturbations in structure that might otherwise go unnoticed in an apparently rigid protein.

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“…From a drug-development perspective, there may be distinct advantages to targeting domains other than SH2, a large domain family present throughout the genome. [25–27] …”

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confidence: 99%

Rhodium(II) Proximity‐Labeling Identifies a Novel Target Site on STAT3 for Inhibitors with Potent Anti‐Leukemia Activity

et al. 2015

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Nearly 40% of children with acute myeloid leukemia (AML) suffer relapse due to chemoresistance, often involving upregulation of the oncoprotein STAT3 (signal transducer and activator of transcription 3). In this paper, rhodium(II)-catalyzed, proximity-driven modification identifies the STAT3 coiled-coil domain (CCD) as a novel ligand-binding site, and we describe a new naphthalene sulfonamide inhibitor that targets the CCD, blocks STAT3 function, and halts its disease-promoting effects in vitro, in tumor growth models, and in a leukemia mouse model, validating this new therapeutic target for resistant AML.

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Structure, dynamics, and binding thermodynamics of the v‐Src SH2 domain: Implications for drug design

Cited by 22 publications

References 62 publications

Molecular Mechanism of Selective Recruitment of Syk Kinases by the Membrane Antigen-Receptor Complex

Molecular Mechanism of Selective Recruitment of Syk Kinases by the Membrane Antigen-Receptor Complex

Constraining Binding Hot Spots: NMR and Molecular Dynamics Simulations Provide a Structural Explanation for Enthalpy−Entropy Compensation in SH2−Ligand Binding

Rhodium(II) Proximity‐Labeling Identifies a Novel Target Site on STAT3 for Inhibitors with Potent Anti‐Leukemia Activity

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