Structural motifs in the RGS RZ subfamily combine to attenuate interactions with Gα subunits

Salem-Mansour, Denise; Asli, Ali; Avital-Shacham, Meirav; Kosloff, Mickey

doi:10.1016/j.bbrc.2018.08.033

Cited by 10 publications

(10 citation statements)

References 30 publications

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“…To map the individual residues that contribute to the interactions of Gα q with each of its partners, we characterized the five complexes detailed above using an energy-based computational methodology developed previously by our lab 45–47,53–55 . The FDPB method was used to calculate the net electrostatic and polar contributions (ΔΔG elec ) of each residue within 15 Å of the Gα q -partner interface in each complex.…”

Section: Resultsmentioning

confidence: 99%

“…We followed the methodology described previously 45–47,53–55,64 to analyze the per-residue contributions of Gα q residues to interactions with their partners (PLC-β3, p63RhoGEF, RGS2/8, GRK2) in the crystal structures mentioned above. The Finite Difference Poisson–Boltzmann (FDPB) method, as implemented in DelPhi 65 , was used to calculate the net electrostatic and polar contributions (ΔΔG elec ) of each residue found within 15 Å of the dimer interface.…”

Section: Methodsmentioning

confidence: 99%

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Structural design principles that underlie the multi-specific interactions of Gαq with dissimilar partners

Navot

Kosloff

2019

Sci Rep

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Gα q is a ubiquitous molecular switch that activates the effectors phospholipase-C-β3 (PLC-β3) and Rho guanine-nucleotide exchange factors. Gα q is inactivated by regulators of G protein signaling proteins, as well as by PLC-β3. Gα q further interacts with G protein-coupled receptor kinase 2 (GRK2), although the functional role of this interaction is debated. While X-ray structures of Gα q bound to representatives of these partners have revealed details of their interactions, the mechanistic basis for differential Gα q interactions with multiple partners (i.e., Gα q multi-specificity) has not been elucidated at the individual residue resolution. Here, we map the structural determinants of Gα q multi-specificity using structure-based energy calculations. We delineate regions that specifically interact with GTPase Activating Proteins (GAPs) and residues that exclusively contribute to effector interactions, showing that only the Gα q “Switch II” region interacts with all partners. Our analysis further suggests that Gα q -GRK2 interactions are consistent with GRK2 functioning as an effector, rather than a GAP. Our multi-specificity analysis pinpoints Gα q residues that uniquely contribute to interactions with particular partners, enabling precise manipulation of these cascades. As such, we dissect the molecular basis of Gα q function as a central signaling hub, which can be used to target Gα q -mediated signaling in therapeutic interventions.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Structural design principles that underlie the multi-specific interactions of Gαq with dissimilar partners

Navot

Kosloff

2019

Sci Rep

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“…We followed the methodology described previously to analyze the per‐residue contributions of Ang2 and Tie2 residues to intermolecular interactions in the complex (Supporting Information Figure S1). The finite difference Poisson‐Boltzmann (FDPB) method was used to calculate the net electrostatic and polar contributions (ΔΔG elec ) of each residue that is within 15 Å of the dimer interface.…”

Section: Methodsmentioning

confidence: 99%

“…We followed the methodology described previously 32,[36][37][38][39][40] to analyze the per-residue contributions of Ang2 and Tie2 residues to intermolecular interactions in the complex (Supporting Information Figure S1).…”

Section: Energy Calculations To Identify Residues That Contribute Dmentioning

confidence: 99%

Residue‐level determinants of angiopoietin‐2 interactions with its receptor Tie2

et al. 2018

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We combined computational and experimental methods to interrogate the binding determinants of angiopoietin‐2 (Ang2) to its receptor tyrosine kinase (RTK) Tie2—a central signaling system in angiogenesis, inflammation, and tumorigenesis. We used physics‐based electrostatic and surface‐area calculations to identify the subset of interfacial Ang2 and Tie2 residues that can affect binding directly. Using random and site‐directed mutagenesis and yeast surface display (YSD), we validated these predictions and identified additional Ang2 positions that affected receptor binding. We then used burial‐based calculations to classify the larger set of Ang2 residues that are buried in the Ang2 core, whose mutations can perturb the Ang2 structure and thereby affect interactions with Tie2 indirectly. Our analysis showed that the Ang2‐Tie2 interface is dominated by nonpolar contributions, with only three Ang2 and two Tie2 residues that contribute electrostatically to intermolecular interactions. Individual interfacial residues contributed only moderately to binding, suggesting that engineering of this interface will require multiple mutations to reach major effects. Conversely, substitutions in substantially buried Ang2 residues were more prevalent in our experimental screen, reduced binding substantially, and are therefore more likely to have a deleterious effect that might contribute to oncogenesis. Computational analysis of additional RTK‐ligand complexes, c‐Kit‐SCF and M‐CSF‐c‐FMS, and comparison to previous YSD results, further show the utility of our combined methodology.

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“…To map the individual residues that contribute to colicin/pyocin-immunity protein interactions, we analyzed the representative X-ray structures of colicins bound to their immunity proteins, using an energybased computational methodology developed previously by our lab [55][56][57][58][59][60][61] . As described in the "Materials and methods" sectio, we calculate the net electrostatic/polar contributions (ΔΔG elec ) of each residue ≤ 15 Å of the colicin-immunity protein interfaces.…”

Section: Residue-level Mapping Of the Interactions Of Colicins With Tmentioning

confidence: 99%

Structural design principles for specific ultra-high affinity interactions between colicins/pyocins and immunity proteins

Shushan

Kosloff

2021

Sci Rep

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The interactions of the antibiotic proteins colicins/pyocins with immunity proteins is a seminal model system for studying protein–protein interactions and specificity. Yet, a precise and quantitative determination of which structural elements and residues determine their binding affinity and specificity is still lacking. Here, we used comparative structure-based energy calculations to map residues that substantially contribute to interactions across native and engineered complexes of colicins/pyocins and immunity proteins. We show that the immunity protein α1–α2 motif is a unique structurally-dissimilar element that restricts interaction specificity towards all colicins/pyocins, in both engineered and native complexes. This motif combines with a diverse and extensive array of electrostatic/polar interactions that enable the exquisite specificity that characterizes these interactions while achieving ultra-high affinity. Surprisingly, the divergence of these contributing colicin residues is reciprocal to residue conservation in immunity proteins. The structurally-dissimilar immunity protein α1–α2 motif is recognized by divergent colicins similarly, while the conserved immunity protein α3 helix interacts with diverse colicin residues. Electrostatics thus plays a key role in setting interaction specificity across all colicins and immunity proteins. Our analysis and resulting residue-level maps illuminate the molecular basis for these protein–protein interactions, with implications for drug development and rational engineering of these interfaces.

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Structural motifs in the RGS RZ subfamily combine to attenuate interactions with Gα subunits

Cited by 10 publications

References 30 publications

Structural design principles that underlie the multi-specific interactions of Gαq with dissimilar partners

Structural design principles that underlie the multi-specific interactions of Gαq with dissimilar partners

Residue‐level determinants of angiopoietin‐2 interactions with its receptor Tie2

Structural design principles for specific ultra-high affinity interactions between colicins/pyocins and immunity proteins

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