Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

HDX-MS of the human eEF1B complex

ABSTRACT: Protein synthesis in eukaryotic cell is spatially and structurally compartmentalized that ensures high efficiency of this process. One of the distinctive features of higher eukaryotes is the existence of stable multi-protein complexes involved in translation. Here we present the data on human guanine-nucleotide exchange factor (GEF) complex, eEF1B, comprising α, β and γ subunits. The project concerns HDX-MS measurements of individual subunits of this complex contributing to understanding of their spatial organization. In addition, we present HDX-MS data on binary complexes of the subunits and on the eEF1B complex as a whole, helping to position the subunits inside the complex.

INSTRUMENT(S): Synapt MS

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Michał Kistowski

LAB HEAD: Prof. Michał Dadlez

PROVIDER: PXD031783 | Pride | 2022-08-02

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	Tania_161107_1Bg_1min.raw.zip	Raw
	Tania_161107_2eEF1BBg_KSCN_1min.raw.zip	Raw
	Tania_161107_3eEF1Bg_KSCN_1min.raw.zip	Raw
	Tania_161107_Bb_norm_W.raw.zip	Raw
	Tania_161107_Bb_norm_W_20161107182828.zip	Other

Items per page:

1 - 5 of 132

Similar Datasets

Biochemical and structural insights into SARS-CoV-2 polyprotein processing by Mpro (HDX-MS pulse labeling)

Project description:SARS-CoV-2, a human coronavirus, is the causative agent of the COVID-19 pandemic, entailing enormous disruption worldwide. Its ~30 kb RNA genome is translated into two large polyproteins subsequently cleaved by viral papain-like protease and main protease (Mpro/nsp5). Polyprotein processing is essential yet incompletely understood. We studied Mpro-mediated processing of the nsp7-10/11 polyprotein, whose mature products are cofactors of the viral replicase, identifying the order of cleavages: 1) nsp9-10, 2) nsp8-9/nsp10-11, and 3) nsp7-8. Integrative modeling based on mass spectrometry (including hydrogen-deuterium exchange and cross-linking) and X-ray scattering yielded three-dimensional models of the nsp7-10/11 polyprotein, and the data suggest that the nsp7-10/11 structure in complex with Mpro strongly resembles the unbound polyprotein. Our array of techniques supports the notion that interplay between polyprotein conformation and junction accessibility determine the preference and order of cleavages. Finally, we leveraged assays of limited proteolysis by Mpro of nsp7-11 using a series of inhibitors/binders to characterize Mpro inhibition using a polyprotein substrate.

2022-12-19 | PXD033702 | Pride

Selective modulation of nuclear receptor LRH-1 through target-ing buried and surface regions of the binding pocket

Project description:Liver receptor homolog-1 (LRH-1) is a phospholipid-sensing nuclear receptor that has shown promise as a target for alleviat-ing intestinal inflammation and disease states characterized by metabolic dysregulation in the liver. LRH-1 contains an unu-sually large ligand binding pocket, and generating synthetic modulators has been challenging. Leveraging a hexahydropen-talene (6HP) core, we have had recent success in generating potent and efficacious agonists through two distinct strategies. We targeted residues deep within the pocket to enhance compound binding, while residues at the mouth of the pocket were targeted to mimic interactions made by endogenous phospholipids. Here, we unite these two designs into one hybrid mole-cule that is the most potent LRH-1 agonist to date. Through a combination of global transcriptomic, biochemical, and struc-tural studies, we show that selective modulation can be driven through contacting deep vs. surface polar regions in the pock-et. While deep pocket contacts convey high-affinity, contacts with the pocket mouth dominate allostery and provide a phos-pholipid-like transcriptional response in cultured cells.

2022-05-04 | PXD031099 | Pride

Biochemical and structural insights into SARS-CoV-2 polyprotein processing by Mpro (HDX-MS continuous labeling)

2022-12-19 | PXD033698 | Pride

Recruitment of trimeric eIF2 by phosphatase non-catalytic subunit PPP1R15B

Project description:HDX-MS of R15B and it's interactions with trimeric eIF2 substrate. Here we integrated diverse approaches to elucidate how the PP1 non-catalytic subu-nit PPP1R15B (R15B) captures its full trimeric eIF2 substrate. We found that the substrate recruitment module of R15B is largely disordered with three short helical elements, H1, H2 and H3. H1 and H2 form a clamp that grasps the substrate in a re-gion remote from the phosphorylated residue. A homozygous N423D variant, adja-cent to H1, reducing substrate binding and dephosphorylation was discovered in a ra-re syndrome with microcephaly, development delay and intellectual disability. These findings explain how R15B captures its 125 kDa substrate by binding the far end of the complex relative to the phospho-site to present it for dephosphorylation by PP1, a paradigm of broad relevance.

2024-05-22 | PXD047538 | Pride

Prion-like low complexity regions enable avid virus-host interactions during HIV-1 infection

Project description:Cellular proteins CPSF6, NUP153 and SEC24C play crucial roles in HIV-1 infection. While weak interactions of short FG peptides to isolated capsid hexamers have been characterized, how these proteins engage biologically relevant extended HIV-1 capsid lattices is unknown. We have identified that prion-like low complexity regions (LCR) enable avid binding of CPSF6, NUP153 and SEC24C to curved hexameric capsid lattices. Structural studies reveal that LCR-LCR interactions mediate multivalent CPSF6 assembly, which are templated by binding of CPSF6 FG peptides to a subset of hydrophobic capsid pockets positioned along adjoining hexamers. CPSF6 LCR mediated avid binding to HIV-1 cores is essential for functional virus-host interactions in infected cells. Investigational drug lenacapavir can access unoccupied hydrophobic pockets in the CPSF6-HIV-1 complex and potently impair HIV-1 inside the nucleus without displacing the tightly bound cellular cofactor from virus cores. These results elucidate previously undescribed mechanisms of virus-host interactions and potent inhibition of HIV-1.

2022-08-31 | PXD030332 | Pride

HDX-MS of TRAPPII reveals a conserved Rab binding interface

Project description:We used hydrogen deuterium exchange mass spectrometry (HDX-MS) to define the interface of TRAPPII with three Rab GTPases (Rab1, Rab11, Rab43). These experiments were performed in the presence of EDTA to generate a nucleotide free stabilised Rab-GEF complex. Significant decreases in deuterium incorporation (defined as >4%, >0.4Da, two tailed T-test p value <0.01) were identified in the TRAPPC2L, TRAPPC4, and TRAPPC5 in the presence of Rab GTPases with no other changes in other TRAPP subunits. Here we show that all three Rab GTPases bind at a conserved interface of TRAPPII.

2020-12-11 | PXD020890 | Pride

Structure of the phosphoinositide 3-kinase p110-p101 complex reveals molecular mechanism of GPCR activation

Project description:The class IB phosphoinositide 3-kinase (PI3K), PI3K, is a master regulator of immune cell function, and is a promising drug target for both inflammatory diseases and cancer. Critical to PI3K function is the association of the p110 catalytic subunit to either a p101 or p84 regulatory subunit, which mediates regulation by G-protein coupled receptors (GPCRs). Here, we report the first structure of a heterodimeric PI3K complex, p110-p101. This structure revealed a unique mode of assembly of catalytic and regulatory subunits distinct from that of other class I PI3K complexes. Multiple oncogenic mutations mapped to these novel interfaces led to increased activation by G. p101 mediates activation through its G binding domain, recruiting the complex to the membrane and allowing for engagement of a secondary site in p110. A nanobody that specifically binds to this p101-G interface blocks activation providing a novel tool to study p101-specific signaling events in vivo.

2021-09-22 | PXD025209 | Pride

Conformational plasticity of RAS Q61 family of neoepitopes results in distinct features for targeted recognition.

Project description:We utilize hydrogen deuterium exchange mass spectrometry (HDX-MS) to probe the solvation and conformational dynamics of HLA-A*01:01/hβ2m complexes that are emptied or loaded with NRAS Q61K peptide, as well as wild-type (Q61) and three additional, common neoepitopes (Q61H, Q61L, Q61R).

2023-11-30 | PXD044838 | Pride

The sequence-specific peptide-binding activity of the protein sulfide isomerase AGR2 directs its’ stable binding to the oncogenic receptor EpCAM

Project description:AGR2 is an oncogenic endoplasmic reticulum (ER)-resident protein disulfide isomerase. AGR2 protein has a relatively unique property for a chaperone in that it can bind sequence-specifically to a peptide motif (TTIYY). A synthetic TTIYY-containing peptide column can be used to affinity-purify AGR2 from crude lysates highlighting peptide selectivity in complex mixtures. Hydrogen-deuterium exchange mass spectrometry localized the dominant region in AGR2 that interacts with the TTIYY peptide to within a structural loop from amino acids 131-135 (VDPSL). A peptide binding site consensus of Tx[IL][YF][YF] was developed for AGR2 by measuring its activity against a alanine mutagenized synthetic peptide library. Screening the human proteome for proteins harboring this consensus motif revealed an enrichment in transmembrane proteins and we focus on validating EpCAM as one such oncogenic protein. Recombinant AGR2 and EpCAM proteins formed a dose-dependent protein-protein interaction in vitro. Proximity ligation assays demonstrated that endogenous AGR2 and EpCAM protein associate in cells. Introducing a single alanine mutation in EpCAM at Tyr251 attenuated its binding to AGR2 in vitro and in cells. Hydrogen-deuterium exchange mass spectrometry was used to identify a stable binding site for AGR2 on EpCAM, adjacent to the TILYY motif and surrounding EpCAM’s detergent binding site. Together, these data define a dominant peptide-binding site on AGR2 that mediates its specific peptide-binding function. A model client protein, EpCAM, is proposed for AGR2 to study how an ER-resident chaperone can dock specifically and regulate assembly of a protein destined for the secretory pathway.

2018-01-22 | PXD005782 | Pride

ATP-competitive and allosteric inhibitors induce differential conformational changes at the autoinhibitory interface of Akt

Project description:We use HDX-MS to interrogate the AKT1 DrLink conformational changes upon binding AKT1 active site inhibitors A-443654, Capivasertib, and Uprosertib, Akt1 allosteric inhibitor MK-2206, and ADP.

2024-08-23 | PXD039028 | Pride