The Crystal Structure of a c-Src Complex in an Active Conformation Suggests Possible Steps in c-Src Activation

Cowan‐Jacob, Sandra W.; Fendrich, Gabriele; Manley, Paul W.; Jahnke, Wolfgang; Fabbro, Doriano; Liebetanz, Janis; Meyer, Thomas

doi:10.1016/j.str.2005.03.012

Cited by 301 publications

(353 citation statements)

References 47 publications

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“…Activation of both Src and Abl is enabled by the disengagement or reconfiguration of the intramolecular interactions just described (7,11,12). Regulation can therefore be thought as a reversible equilibrium whereby the SH3, SH2, and catalytic domains are either dissociated or self-assembled in one or more configurations.…”

Section: Significancementioning

confidence: 99%

Two-state dynamics of the SH3–SH2 tandem of Abl kinase and the allosteric role of the N-cap

Corbi‐Verge

Marinelli

Zafra-Ruano

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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The regulation and localization of signaling enzymes is often mediated by accessory modular domains, which frequently function in tandems. The ability of these tandems to adopt multiple conformations is as important for proper regulation as the individual domain specificity. A paradigmatic example is Abl, a ubiquitous tyrosine kinase of significant pharmacological interest. SH3 and SH2 domains inhibit Abl by assembling onto the catalytic domain, allosterically clamping it in an inactive state. We investigate the dynamics of this SH3-SH2 tandem, using microsecond all-atom simulations and differential scanning calorimetry. Our results indicate that the Abl tandem is a two-state switch, alternating between the conformation observed in the structure of the autoinhibited enzyme and another configuration that is consistent with existing scattering data for an activated form. Intriguingly, we find that the latter is the most probable when the tandem is disengaged from the catalytic domain. Nevertheless, an amino acid stretch preceding the SH3 domain, the so-called N-cap, reshapes the free-energy landscape of the tandem and favors the interaction of this domain with the SH2-kinase linker, an intermediate step necessary for assembly of the autoinhibited complex. This allosteric effect arises from interactions between N-cap and the SH2 domain and SH3-SH2 connector, which involve a phosphorylation site. We also show that the SH3-SH2 connector plays a determinant role in the assembly equilibrium of Abl, because mutations thereof hinder the engagement of the SH2-kinase linker. These results provide a thermodynamic rationale for the involvement of N-cap and SH3-SH2 connector in Abl regulation and expand our understanding of the principles of modular domain organization.allosteric inhibitors | protein-protein interactions | macromolecular assembly | population shift | leukemia

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Section: Significancementioning

confidence: 99%

Two-state dynamics of the SH3–SH2 tandem of Abl kinase and the allosteric role of the N-cap

Corbi‐Verge

Marinelli

Zafra-Ruano

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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“…Crystal structures (3,4) and mutational studies complemented by molecular dynamics (MD) simulations (9) are consistent with this view. However, one crystal structure of c-Src in a partially activated state in which the SH2-SH3 modules are reassembled in a different orientation with respect to the catalytic domain (10), suggests that the situation could be considerably more complex. In this regard, it seems likely that different signals could promote different assembly states, leading to differently configured activated kinases.…”

Section: Coarse-grained Model | Bayesian Analysis | Folding | Saxs | mentioning

confidence: 99%

Multidomain assembled states of Hck tyrosine kinase in solution

Yang

Blachowicz

Makowski

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

194

199

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An approach combining small-angle X-ray solution scattering (SAXS) data with coarse-grained (CG) simulations is developed to characterize the assembly states of Hck, a member of the Srcfamily kinases, under various conditions in solution. First, a basis set comprising a small number of assembly states is generated from extensive CG simulations. Second, a theoretical SAXS profile for each state in the basis set is computed by using the Fast-SAXS method. Finally, the relative population of the different assembly states is determined via a Bayesian-based Monte Carlo procedure seeking to optimize the theoretical scattering profiles against experimental SAXS data. The study establishes the concept of basis-set supported SAXS (BSS-SAXS) reconstruction combining computational and experimental techniques. Here, BSS-SAXS reconstruction is used to reveal the structural organization of Hck in solution and the different shifts in the equilibrium population of assembly states upon the binding of different signaling peptides.T yrosine kinases of the Src-family are large multidomain allosteric enzymes implicated in the signaling pathways regulating cell growth and proliferation (1). The key role that Src kinases play in the onset of many human diseases, particularly cancer, makes them important targets for therapeutic intervention (2).All Src kinases share a common structural organization comprising the SH3 and SH2 binding domains followed by a highly conserved catalytic domain connected by flexible linkers (1). Crystal structures have offered a detailed view of the down-regulated forms of Hck and c-Src (3, 4), characterized by a compact assembled form stabilized by auto-inhibitory intramolecular interactions between the N and C terminus of the catalytic domain with the SH3 and SH2 modules, respectively. The SH2 and SH3 modules of the Src-family tyrosine kinases play dual roles: both are involved in the auto-inhibitory intramolecular interactions down-regulating kinase activity, but can also serve as possible binding receptors for cellular signals leading to kinase activation.In a broadly accepted paradigm, Src inactivation/activation is pictured in terms of a simple two-state process involving an assembled (inactive) and a disassembled (active) conformation, i.e., once any of the intramolecular interaction is released, the kinase domain switches to a disassembled catalytically active state targeting down-stream cellular substrates (5-8). Crystal structures (3, 4) and mutational studies complemented by molecular dynamics (MD) simulations (9) are consistent with this view. However, one crystal structure of c-Src in a partially activated state in which the SH2-SH3 modules are reassembled in a different orientation with respect to the catalytic domain (10), suggests that the situation could be considerably more complex. In this regard, it seems likely that different signals could promote different assembly states, leading to differently configured activated kinases. While it is understood that Src increases its activity in respon...

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“…The addition of 1 or more phosphates to hydroxyl-bearing amino acids is catalyzed by protein kinases, a large family of enzymes that all share a wellconserved catalytic core of Ϸ250 residues (13). Despite the simplicity of the reaction catalyzed by these enzymes, the protein kinases are regulated in complex ways that may involve phosphorylation by other kinases (14), membrane and organelle localization through scaffolding proteins (15), and protein-protein and domain-domain interactions through regulator modules (16,17). In the last decade, a full appreciation for the conformational complexity of these enzymes has been developed (11,18).…”

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confidence: 99%