IndraLab
Statements
sparser
"To test the extent to which the formation of Grb2-Sos1 complex with a 2:1 stoichiometry may be influenced by steric hindrance, we next measured the binding of full-length Grb2 to various single, double and triple mutant constructs of the PR domain, mutated with respect to one or more of the S1–S4 sites ( xref and xref )."
sparser
"Based on the concentrations used for these studies, it is estimated that the affinity of SOS1 scrambled for GRB2 is greater than 50 µM. These experiments suggest that the PxxPxR motifs or sequences needed for interactions with these motifs in the context of short peptides are not required for the high-affinity binding of GRB2 to SOS1."
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"Close examination of the relative physical location of S1-S4 sites within the PR domain suggests that the length of the intervening linker spanning various pairwise combinations of these sites is likely to play a key role in determining the stoichiometry of Grb2 and Sos1 complex (XREF_FIG)."
sparser
"Since these observations directly challenges the current model of SH3 domain binding reactions, we further examined the requirement of PxxPxR motifs for high affinity GRB2:SOS1 binding using dynamic light scattering, which determines the hydrodynamic radius of a protein complex."
sparser
"However, these receptors also bind adaptor molecules which form complexes with other signalling molecules, such as the regulatory subunit p85 of the phosphatidylinositol 3′-kinase (PI3K), and GRB2, which binds the nucleotide exchange molecule SOS1, activating RAS and the ERK MAP-kinase pathway."
reach
"This Ras dependent MAP kinase activation requires tyrosine phosphorylation of " docking proteins, " including the shc adapter protein, and depends upon recruitment of Grb2 and Sos1 complexes to the plasma membrane, thus resembling the pathway of MAP kinase activation employed by the receptor tyrosine kinases."
sparser
"Alternatively, they are adaptor molecules forming complexes with enzymes, such as Grb2, which forms a complex with SOS1, a nucleotide exchange molecule that activates Ras, and the p85 regulatory subunit of phosphatidylinositol 3'-kinase (PI3-kinase), which forms a complex with the p110 catalytic subunit."
sparser
"Herein, using an array of biophysical methods, we provide evidence that although the Grb2 adaptor can potentially bind to all four PXψPXR motifs (designated herein S1-S4) located within the Sos1 guanine nucleotide exchange factor, the formation of the Grb2-Sos1 signaling complex occurs with a 2:1 stoichiometry."
sparser
"However, owing to the advanced inventory of GRAP2 and GRB2 interactors, such excess is more likely due to the competition that exists between the GRAP2–SLP-76, GRB2–SHIP1, GRB2–SOS1, and GRB2–THEMIS complexes and the larger pool of free GRAP2 and GRB2 adaptors for binding to phosphorylated LAT molecules."
sparser
"Grb2-Sos1 interaction, mediated by the canonical binding of N-terminal SH3 (nSH3) and C-terminal SH3 (cSH3) domains of Grb2 to proline-rich motifs within Sos1, plays a central role in relaying external signals from receptor tyrosine kinases (RTKs) at the cell surface to downstream effectors and regulators such as Ras within the cytosol ( xref – xref )."
sparser
"Our thermodynamic data suggest that the bivalent binding of Grb2 to Sos1 with a 2:1 stoichiometry is under cooperative control in that the binding of nSH3 domain within one Grb2 molecule to one of the S1–S4 sites within Sos1 facilitates the binding of nSH3 domain within a second Grb2 molecule."
sparser
"The Grb2 interaction with the SOS1 PR domain is at least 100-fold stronger than the interactions of the individual nSH3/cSH3 domains with the truncated SOS1 PR peptides. xref This suggests that Grb2-SOS1 association likely involves simultaneous binding of nSH3 and cSH3 domains to the SOS1 PR domain."
sparser
"Importantly, the presence of multiple binding sites within Sos1 appears to provide a physical route for Grb2 to hop in a flip-flop manner from one site to the next through facilitated diffusion and such rapid exchange forms the basis of positive cooperativity driving the bivalent binding of Grb2 to Sos1 with high affinity."
sparser
"To gain insights into the structural basis of the binding of Grb2 to Sos1 as two independent monomers and to compare this mode of binding to that of a homodimer, we built appropriate structural models of Grb2 bivalently bound to PXψPXR motifs located within the S1 and S4 sites in the PR domain of Sos1 ( xref )."
sparser
"Activated tyrosine kinase receptors can stimulate intracellular-signalling pathways which regulate cell proliferation, such as the Ras-mitogen-activated protein kinase-ERK (Ras-MAPK-ERK) pathway [ xref , xref ]; once phosphorylated, they recruit growth factor receptor bound protein 2 (GRB2) which is constitutively associated with Sos1 protein, a guanine nucleotide exchange factor of the small GTP-ase, Ras."
sparser
"However, integrins and RTKs can also stimulate Ras-mediated activation of the MAPK cascade independently of FAK and do so by binding Grb2-Sos1 directly or indirectly via a Shc protein (van der Geer and Pawson, xref ; Rojas et al., xref ; Wary et al., xref ; Kroll and Waltenberger, xref ; McKay and Morrison, xref )."
reach
"Thus, while the binding of Sos1 to cSH3 domain within the wildtype Grb2 requires the formation of three ion pairs, namely R1156-E171, R1157-D187 and R1158-D190, formation of only the E171-R1156-G173D tripartite salt bridge may be necessary for the binding of Sos1 to the cSH3 domain within the Grb2_G173D construct."
reach
"Our thermodynamic data suggest that the bivalent binding of Grb2 to Sos1 with a 2:1 stoichiometry is under cooperative control in that the binding of nSH3 domain within one Grb2 molecule to one of the S1-S4 sites within Sos1 facilitates the binding of nSH3 domain within a second Grb2 molecule."
sparser
"Moreover, to exclude the possibility that SUMOylation enhancing the formation of Grb2-Sos1 complex is attributed to Sos1 SUMOylation, we investigated whether endogenous Sos1 can be SUMOylated by using in vivo Ni 2+ -NTA pulldown (Figure xref E) and immunoprecipitation (Figure xref F) assays."
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"However, these receptors also bind adaptor molecules which form complexes with other signalling molecules, such as the regulatory subunit p85 of the phosphatidylinositol 3 '-kinase (PI3K), and GRB2, which binds the nucleotide exchange molecule SOS1, activating RAS and the ERK MAP-kinase pathway."
sparser
"Ternary protein complexes
(Shc1:Grb2:SOS1 and Grb2:Gab1:PI3K) were identified, and we found
that EGFR downstream signaling is based on constitutively bound (Grb2:SOS1
and Grb2:Gab1) as well as on agonist-dependent protein associations
with transient interaction properties (Grb2:Shc1 and Grb2:PI3K)."
reach
"Nonetheless, the demonstration that the bivalent binding of Grb2 to Sos1 through a variety of pairwise combinations of S1-S4 sites is feasible strongly argues that conformational heterogeneity may be a hallmark of Grb2-Sos1 signaling complex -- formation of conformationally-distinct complexes through the employment of distinct pairwise combinations of S1-S4 sites."
reach
"To test the extent to which the formation of Grb2 and Sos1 complex with a 2:1 stoichiometry may be influenced by steric hindrance, we next measured the binding of full-length Grb2 to various single, double and triple mutant constructs of the PR domain, mutated with respect to one or more of the S1-S4 sites (XREF_FIG and XREF_TABLE)."
reach
"These data suggest an interesting hypothesis whereby multivalent, cooperative interactions between LAT, Grb2, and Sos1 promote LAT oligomerization, whereas recruitment of PLC-gamma1-GADS-SLP-76 complexes block Grb2-LAT associations, and therefore limit the overall size of LAT oligomers."
sparser
"Herein, using an array of biophysical methods, we provide evidence that although Grb2 adaptor can potentially bind to all four PXψPXR motifs — designated herein S1, S2, S3 and S4 — located within the Sos1 guanine nucleotide exchange factor, the formation of Grb2-Sos1 signaling complex occurs with a 2:1 stoichiometry."
sparser
"Finally, peptides derived from PxxP motifs found in SOS1interact with individual SH3 domains from GRB2 with affinities ranging from 20 µM to 1 mM [ xref – xref ], whereas the association of full-length GRB2 with full-length SOS1 or the complete proline rich regions (PRR) of SOS1 and CBL has affinities of 300–400 nM [ xref – xref ]."
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"The resulting GRB2 (growth factor receptor binding protein) binding (via the SH2 domain) to the phosphorylated receptor and subsequent binding of GRB2 to SOS1 (son of sevenless 1, a guanine nucleotide exchange factor) activates the Ras protein to exchange GDP for GTP [128,129,130,131,132]."
sparser
"Accordingly, it can be proposed that (1) HER2 pYP-bound SH2 potentiates GRB2 SH3 domain interactions with SOS1 (an allosteric mechanism); (2) the SH2 domain blocks cSH3,enabling nSH3 to bind SOS1 first before cSH3 follows (an avidity-based mechanism); and (3) the allosteric behavior of cSH3 to other domains appears to be unidirectional, although there is an allosteric effect between the SH2 and SH3 domains."
sparser
"Given that Sos1 competes with a diverse array of other downstream effectors for binding to Grb2, including the adaptor proteins Gab1 and SLP76 ( xref , xref ), the endocytic GTPase dynamin1 ( xref , xref ), the ubiquitin ligase Cbl ( xref , xref , xref ) and the cell cycle inhibitor p27kip1 ( xref ), the ability of cSH3 domain to dictate the binding of Grb2 to Sos1 in an oriented manner may also play an important role in determining the fraction of Grb2-Sos1 pool."
sparser
"Son of Sevenless homolog 1 (Sos1), another GEF, is also implicated in Rac1 activation in diabetic milieu; a regulator of Sos1, 66kDa proto-oncogene Src homologous-collagen homologue (p66Shc), regulates the of binding of Sos1 with the growth factor receptor-bound protein 2 (Grb2) [ xref ], and in diabetes, the binding of Sos1 with Grb2 is decreased, resulting in Rac1 activation [ xref ]."
sparser
"In these regards, one potential combinatorial drug candidate would be BI 1701963, a clinical pan-Ras inhibitor provided that wild-type Ras proteins are involved in the resistance to K-Ras G12C inhibition. xref Furthermore, co-targeting K-Ras G12C with SOS1 or GRB2 could also be promising in clinic given that interactions between SOS1 and K-Ras, as well as SOS1 with its adaptor GRB2 are required for RTK-mediated adaptive Ras reactivation. xref , xref , xref Currently, drugs targeting GRB2-SOS1 interaction are under investigation. xref Synergistic antiproliferative activity has been reported upon co-inhibition of K-Ras G12C and SOS1, although these findings need to be validated in in vivo studies. xref In addition, future studies are expected to evaluate the combinatorial effects of K-Ras G12C and GRB2 inhibition."
"To gain further insight into the importance of Grb2 for CML, peptides that disrupt Grb2-SoS complexes were tested. These high-affinity Grb2-binding peptides (HAGBPs) can autonomously shuttle into cells and function by binding to the N-terminal SH3 domain of Grb2. The HAGBPs were analyzed for their effects on Bcr-Abl-expressing cell lines and freshly isolated CML blast cells from patients. They induced a dramatic decrease in the proliferation of CML cell"
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"In light of previous studies, the most straightforward interpretation of this finding is that Grb2 binds in a bivalent manner to Sos1 to form the Grb2 and Sos1 complex with a 2:1 stoichiometry and that this interaction is mediated through the binding of nSH3 domain within each Grb2 molecule to only two out of a possible four available S1-S4 sites within Sos1."
reach
"These observations fit well with previous studies showing that SOS1 peptides bind to individual SH3 domains from GRB2 with affinities ranging from 20 microM to 1 mM [XREF_BIBR - XREF_BIBR], while the affinity for the binding of full-length GRB2 with full-length SOS1 or the complete PRR of SOS1 and CBL is between 300 and 400 nM (XREF_TABLE) and [XREF_BIBR - XREF_BIBR]."
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"Given that Sos1 competes with a diverse array of other downstream effectors for binding to Grb2, including the adaptor proteins Gab1 and SLP76, the endocytic GTPase dynamin1, the ubiquitin ligase Cbl and the cell cycle inhibitor p27kip1, the ability of cSH3 domain to dictate the binding of Grb2 to Sos1 in an oriented manner may also play an important role in determining the fraction of Grb2-Sos1 pool."
sparser
"Grb2 forms a complex with Sos1 through interactions of its C- and N-terminal SH3 domains with “P-x-x-P-x-R” sequences in Sos1, with the N-terminal SH3 domain-binding affinity of WT peptide 1 having been reported as K d = 39 μM by isothermal calorimetry. xref In our current work, in vitro Grb2 SH3 N-domain binding affinities of synthetic peptides were determined using a Trp-fluorescence assay. xref The WT peptide 1 , as well as the open-chain metathesis precursors 5a – 5c , showed similar binding affinities (K d ≈ 11 – 14 μM; xref )."
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"ERK1/2 have long been known to phosphorylate SOS1 at four critical residues reducing the binding of SOS1 to GRB2 [XREF_BIBR], the effect of which may be to diffuse signaling away from the receptor complex even if SOS1 's membrane binding domains retain SOS1 near lipid products at the membrane."
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"To gain insights into the structural basis of the binding of Grb2 to Sos1 as two independent monomers and to compare this mode of binding to that of a homodimer, we built appropriate structural models of Grb2 bivalently bound to PXpsiPXR motifs located within the S1 and S4 sites in the PR domain of Sos1 (XREF_FIG)."
sparser
"Here, we demonstrate that (a) Sos-1, E3b1, and Eps8 assemble into a tricomplex in vivo under physiological conditions; (b) Grb2 and E3b1 bind through their SH3 domains to the same binding site on Sos-1, thus determining the formation of either a Sos-1–Grb2 (S/G) or a Sos-1–E3b1–Eps8 (S/E/E8) complex, endowed with Ras- and Rac-specific GEF activities, respectively; (c) the Sos-1–Grb2 complex is disrupted upon RTKs activation, whereas the S/E/E8 complex is not; and (d) in keeping with the previous result, the activation of Ras by growth factors is short-lived, whereas the activation of Rac is sustained."
sparser
"To elucidate whether the cSH3 domain is auto-inhibited for binding to Sos1 in the context of full-length wildtype Grb2 (Grb2_WT) or whether the binding of Sos1 to the nSH3 domain is required for its inhibition, we measured the binding of Sos1 peptide to D15G mutant of full-length Grb2 (Grb2_D15G)."
sparser
"On the contrary, it is highly plausible that the binding of PR domain through one of its four S1–S4 sites to the nSH3 domain of one monomer within Grb2 homodimer sterically blocks the binding of second Grb2 monomer to another PR domain so as to prevent the formation of Grb2-Sos1 complex with a 2:2 stoichiometry."
"Tyrosine phosphorylation led to the association of BCR with the RAS guanine nucleotide exchange complex GRB2-SOS in vivo via the GRB2 SH2 domain, linking BCR to RAS signaling (Maru, Y., Peters, K. L., Afar, D. E. H., Shibuya, M., Witte, O. N., and Smithgall, T. E. (1995) Mol. Cell. Biol. 15, 835-842). In the present study, we demonstrate that BCR Tyr-246 and at least one of the closely spaced tyrosine residues, Tyr-279, Tyr-283, and Tyr-289 (3Y cluster), are phosphorylated by FES both in vitro and in 32Pi-labeled cells. Mutagenesis of BCR Tyr-177 to Phe completely abolished FES-induced BCR binding to the GRB2 SH2 domain, identifying Tyr-177 as an additional phosphorylation site for FES."
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"The fact that not only Sos1_PR domain but also the Sos1_S1 peptide inhibits the binding of Gab1_PR domain to Grb2 further cements our thesis that Sos1 and Gab1 bind to Grb2 in a mutually exlusive manner via steric hindrance and allosteric conformational changes so as to alter the structural configuration of the binding grooves within each other 's binding sites within Grb2 upon association."
sparser
"As shown in, Ras can be actively copatterned to
bait-PAR-Grb2 enriched areas, however, the majority of analyzed cells
showed a homogenous HRas-CFP membrane distribution, indicating that
Grb2:SOS1 or SOS1 alone dissociates from the receptor complex to activate
Ras at the plasma membrane."
reach
"Here, we demonstrate that (a) Sos-1, E3b1, and Eps8 assemble into a tricomplex in vivo under physiological conditions; (b) Grb2 and E3b1 bind through their SH3 domains to the same binding site on Sos-1, thus determining the formation of either a Sos-1-Grb2 (S/G) or a Sos-1-E3b1-Eps8 (S/E/E8) complex, endowed with Ras- and Rac specific GEF activities, respectively; (c) the Sos-1 and Grb2 complex is disrupted upon RTKs activation, whereas the S/E/E8 complex is not; and (d) in keeping with the previous result, the activation of Ras by growth factors is short lived, whereas the activation of Rac is sustained."
sparser
"Nonetheless, the demonstration that the bivalent binding of Grb2 to Sos1 through a variety of pairwise combinations of S1–S4 sites is feasible strongly argues that conformational heterogeneity may be a hallmark of Grb2-Sos1 signaling complex — formation of conformationally-distinct complexes through the employment of distinct pairwise combinations of S1–S4 sites."
sparser
"The resulting GRB2 (growth factor receptor binding protein) binding (via the SH2 domain) to the phosphorylated receptor and subsequent binding of GRB2 to SOS1 (son of sevenless 1, a guanine nucleotide exchange factor) activates the Ras protein to exchange GDP for GTP [ xref , xref , xref , xref , xref ]."
sparser
"Since structural data of the Grb2–SOS1 complex are unavailable, to verify the concurrent nSH3/cSH3–SOS1 associations, we construct the four most probable Grb2–SOS1 complex models and sample the conformational ensembles of complexes by using replica-exchange molecular dynamics (REMD) simulations."
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"Surprisingly, there were no statistically significant differences in the affinity for the interaction of GRB2 with SOS1 proteins with sites 2, 3, Z or 4 deleted alone or together compared to WT SOS1 or DeltaH and stoichiometry for the binding of GRB2 to SOS1 proteins with sites 1, 2, 3, Z or 4 deleted alone or together compared to WT SOS1 (XREF_TABLE and XREF_FIG)."
sparser
"Surprisingly, there were no statistically significant differences in the affinity for the interaction of GRB2 with SOS1 proteins with sites 2, 3, Z or 4 deleted alone or together compared to WT SOS1 or ΔH and stoichiometry for the binding of GRB2 to SOS1 proteins with sites 1, 2, 3, Z or 4 deleted alone or together compared to WT SOS1 ( xref and xref )."
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"Moreover, our results revealed that the enhancement of ERK activities was attributed to the SUMO1 modification of Grb2 facilitating Grb2 binding with Sos1 other than other interacting proteins such as EGFR, SHP2, FAK, PTPalpha and Gab1, which are all considered connecting the ERK activities and cell phenotypes."
reach
"Moreover, when the hyperphosphorylation of the Rafs and Sos1 reached peak levels (0.5-5 microM), EGF induced MEK activation was suppressed, whereas activation of the EGFR and autophosphorylation on Y1068, the site that mediates recruitment of the Grb2 and Sos1 complex, was largely unaffected."
sparser
"To understand stoichiometry and the underlying thermodynamic forces driving the formation of Grb2-Sos1 signaling complex, we next measured the binding of full-length Grb2 to the PR domain of Sos1 using ITC (representative data are shown in xref and all data summarized in xref )."
sparser
"The failure of S1/S2 and S2/S3 pairwise combinations to afford bivalent binding of Grb2 to Sos1 thus most likely results from steric hindrance in that binding of one molecule of Grb2 to S1 site physically blocks the binding of a second molecule of Grb2 to S2 site and vice versa."
reach
"On the contrary, it is highly plausible that the binding of PR domain through one of its four S1-S4 sites to the nSH3 domain of one monomer within Grb2 homodimer sterically blocks the binding of second Grb2 monomer to another PR domain so as to prevent the formation of Grb2 and Sos1 complex with a 2:2 stoichiometry."
reach
"In light of the fact that the much shorter peptide docked at the cSH3 domain within Grb2 is unlikely to sterically hinder the binding of Sos1_PR domain to the neighboring nSH3 domain, we argue that the ability of Gab1 to restrict the binding of Sos1 to Grb2 resides not only in steric hindrance but may also be due to its ability to cause a subtle conformational change within Grb2 upon binding to the cSH3 domain."
sparser
"A large number of these PPIs are mediated by multiple SLiM classes or SLiM instances, for example, in the interaction between GRB2 (uniprot:P62993) and SOS1 (uniprot:Q07889); SOS1 has seven putative SH3 motifs and GRB2 has two SH3 domains, potentially this can equate to 14 binding interfaces for a single PPI."
sparser
"Because both the N-terminal and C-terminal SH3 domains of Grb2 are known to interact with Sos1 ( xref ), the binding of SNTA1 and P66shc would be expected to weaken the Sos1-Grb2 interaction, making Sos1 more available to interact with Eps8-E3b1, which is important for Rac1 activation."
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"Upon the activation of EGFR or other RTKs, Grb2 recruits Sos1 to the membrane to form Grb2 and Sos1 complex, which is crucial for signaling transduction and sequentially leads to the activation of Ras/MEK/MAPK (generally recognized as extracellular signal regulated kinase, ERK) [XREF_BIBR]."
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"Although efforts by others and us directed at the determination of X-ray or NMR structures of full-length Grb2 bound to Sos1 or Gab1 constructs of any sort over the past decade or so have met no success, presumably due to the transient nature of Sos1, Grb2, and Gab1 complex, we have nonetheless made an attempt here to create a crude 3D structural model of how the binding of one molecule of Sos1 to the nSH3 domain may allosterically induce a conformational change within Grb2 such that the loading of a second molecule of Sos1 onto the cSH3 domain is blocked and, in so doing, allowing Gab1 access to the cSH3 domain in an exclusively non competitive manner to generate the Sos1-Grb2-Gab1 ternary complex (XREF_FIG)."
sparser
"Upon the activation of EGFR or other RTKs, Grb2 recruits Sos1 to the membrane to form Grb2-Sos1 complex, which is crucial for signaling transduction and sequentially leads to the activation of Ras/MEK/MAPK (generally recognized as extracellular signal-regulated kinase, ERK) [ xref ]."
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"Mechanistically, Tnk1 and Kos1 can directly tyrosine phosphorylate growth factor receptor binding protein 2 (Grb2), which promotes disruption of the Grb2 and Sos1 complex that mediates growth factor induced Ras activation, providing dynamic regulation of Ras GEF activity with suppression of Ras."
sparser
"In this manner, the binding of cSH3 domain to S1 site could restrict the orientation of Grb2 relative to Sos1, while the binding of nSH3 domain to one of the three available S2–S4 sites may result in the formation of various structurally and conformationally distinct Grb2-Sos1 complexes."
sparser
"Our group recently reported two unrelated chemical series that bind to the same hydrophobic pocket on SOS1 as part of a RAS:SOS1:RAS complex. xref – xref We have previously shown that these compounds activate nucleotide exchange on RAS, resulting in increased levels of cellular RAS-GTP and, at low concentrations, enhance phosphorylation of extracellular regulared kinase (ERK) 1/2 via the RAS-MAPK signaling pathway. xref At higher doses of compound, phosphorylated ERK (pERK) phosphorylates SOS1 leading to disruption of the SOS1-GRB2 complex and dissociation of RAS from SOS1, thereby achieving a compound-mediated negative feedback loop that results in reduced ERK phosphorylation."
sparser
"In light of previous studies ( xref , xref ), the most straightforward interpretation of this finding is that Grb2 binds in a bivalent manner to Sos1 to form the Grb2-Sos1 complex with a 2:1 stoichiometry and that this interaction is mediated through the binding of nSH3 domain within each Grb2 molecule to only two out of a possible four available S1–S4 sites within Sos1."
sparser
"A possible explanation
for the reduced fast diffusion of SOS1 and Gab1 in comparison to Shc1
might be that Gab1 and SOS1 are not existing in isolation, as they
were reported to form stable SOS1:Grb2, Grb2:Gab1, and even SOS1:Grb2:Gab1
ternary complexes with different stoichiometries. xref , xref It is likely, especially in the context of a cellular milieu, that
the measured fast recovery kinetics were not obtained from isolated
Gab1 and SOS1 molecules, but instead from stable macromolecular protein
assemblies, which would diffuse slower than single Shc1 molecules
in the cytosol."
reach
"This equation can be rewritten as : (9) or in other words : (10) where is the effective equilibrium constant for the binding of a Grb2 from solution to a Grb2 and Sos1 complex to form a Grb2, Sos1, and Grb2 complex : (11) To make predictions about the binding of Grb2 to the complete polyproline rich domain of Sos1 (1117-1319) we must estimate the values of the unknown equilibrium constants for the binding of the N- and C-SH3 domains of Grb2 to RP on Sos1."
sparser
"Although our previous studies have shown that the isolated nSH3 domain of Grb2 can potentially bind to peptides derived from all four S1–S4 motifs in a physiologically-relevant manner ( xref , xref , xref ), the precise mechanism of the assembly of Grb2-Sos1 signaling complex remains hitherto poorly understood."
sparser
"To further characterize the Grb2-Sos1 interaction in biophysical terms, we measured the binding of nSH3 and cSH3 domains to peptides derived from S1–S4 sites containing the PXψPXR motifs using ITC. xref and xref show representative data obtained upon conducting such measurements, while corresponding thermodynamic parameters are reported in xref ."
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"However, SHP-2 also positively modulates signaling, e.g. via dephosphorylation of a C-terminal inhibitory phosphorylation site in Src family members, thereby activating them [XREF_BIBR], or by acting as an adaptor for binding of the Grb2 and SOS1 complex, thus promoting Ras activation [XREF_BIBR]."
sparser
"In addition, compared with low-grade PCa, high-grade PCa tissue samples have (1) one more abundant protein complex, (2) increased assembly of 10 protein complexes including the GRB2-SOS1 complex, (3) five less abundant protein complexes, and (4) decreased assembly of 12 complexes including the PPP2CA-PPP2R1A complex."
sparser
"Moreover, our results revealed that the enhancement of ERK activities was attributed to the SUMO1 modification of Grb2 facilitating Grb2 binding with Sos1 other than other interacting proteins such as EGFR, SHP2, FAK, PTPα and Gab1, which are all considered connecting the ERK activities and cell phenotypes."
sparser
"Most efforts focus on blocking Ras regulators and effectors, such as SOS1, B-Raf, and PI3K α . xref – xref The Grb2–SOS1 interaction has attracted increasing attention in the clinic, and recent targeted therapy has shown that synthesized peptide inhibitors can interrupt the Grb2–SOS1 interaction, effectively preventing tumor development. xref – xref "
sparser
"Ras is activated by the guanine
nucleotide exchange factor SOS1 by induction of the exchange of GDP
to GTP. xref So far it is not clear whether
this occurs through dissociation of the Grb2:SOS1 complex from the
receptor and translocation to the membrane-bound Ras, or by active
recruitment of Ras to the activated Grb2:SOS1 complex."
sparser
"Thus, while this intervening linker is comprised of 55–133 residues for the pairwise combinations of S1–S4 sites that favor bivalent binding of Grb2 to Sos1 (S1/S4, S1/S3, S2/S4 and S3/S4), it merely spans 23–26 residues for the pairwise combinations that result in the formation of Grb2-Sos1 complex with a 1:1 stoichiometry (S1/S2 and S2/S3)."
sparser
"Knockdown of Grb2 reduced the ERK activity and suppressed cell motility and tumorigenesis in vitro and in vivo , which were all rescued by stable ectopic re-expression of wild-type Grb2 but not the mutant Grb2 K56R . Furthermore, Grb2 SUMOylation at K 56 increased the formation of Grb2-Sos1 complex, which sequentially leads to the activation of Ras/MEK/MAPK pathway."
sparser
"Among most dramatically dysregulated protein complexes, the GRB2-SOS1 complex has significantly higher (Z score difference = 1.57, q = 0.017) assembly level, whereas the PPP2CA-PPP2R1A complex has significantly lower (Z score difference = − 1.64, q = 0.009) assembly level, in high-grade PCa than in low-grade PCa."
sparser
"The observation that monovalent binding is weaker than bivalent binding when the number of potential sites within the PR constructs is reduced to one but comparable and somewhat stronger than bivalent binding when more than one potential site is available unequivocally implicates the role of facilitated diffusion in mediating Grb2-Sos1 interaction."
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"Several events are speculated : (1) growth factor binds to its tyrosine kinase receptor, which autophosphorylates and recruits Grb2 and Sos1 complexes; (2) at the plasma membrane, the activated receptor and signaling complexes initiate the first phase of Ras and MAPK signaling; (3) alpha-adaptin and additional endocytic adaptors such as Dab2 mediate assembly of clathrin coated pits and the formation of endosomes; 4) the endosome undergoes disassembly of adaptin and clathrin coating, and at the same time, the cargo proteins undergo phosphorylation triggered rearrangement."
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"Thus, while this intervening linker is comprised of 55-133 residues for the pairwise combinations of S1-S4 sites that favor bivalent binding of Grb2 to Sos1 (S1/S4, S1/S3, S2/S4 and S3/S4), it merely spans 23-26 residues for the pairwise combinations that result in the formation of Grb2 and Sos1 complex with a 1:1 stoichiometry (S1/S2 and S2/S3)."
sparser
"Although the exact biological function of SHP2 is not clear, it is known that SHP2 acts as key node linking RTK–RAS signalling, probably acting as a scaffold protein which binds GRB2 and SOS1, therefore promoting the activation of RAS proteins by stimulating GDP for GTP exchange [ xref , xref ]."
sparser
"However, SHP-2 also positively modulates signaling, e.g. via dephosphorylation of a C-terminal inhibitory phosphorylation site in Src family members, thereby activating them
[ xref ], or by acting as an adaptor for binding of the Grb2-SOS1 complex, thus promoting Ras activation
[ xref ]."
reach
"Since structural data of the Grb2–SOS1 complex are unavailable, to verify the concurrent nSH3/cSH3–SOS1 associations, we construct the four most probable Grb2–SOS1 complex models and sample the conformational ensembles of complexes by using replica-exchange molecular dynamics (REMD) simulations."
reach
"Importantly, the presence of multiple binding sites within Sos1 appears to provide a physical route for Grb2 to hop in a flip-flop manner from one site to the next through facilitated diffusion and such rapid exchange forms the basis of positive cooperativity driving the bivalent binding of Grb2 to Sos1 with high affinity."
sparser
"In an attempt to test the validity of our structural models and to shed light on macromolecular dynamics underlying the assembly of Grb2-Sos1 signaling complex, we performed MD simulations on structural models of Grb2 bivalently bound to PXψPXR motifs located within the S1 and S4 sites in the PR domain of Sos1 either as two independent monomers (Grb2-PR-Grb2), or alternatively as a homodimer ([Grb2] 2 -PR) ( xref )."