Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

A Novel Mechanism Mediated by Allosteric Disulfide Bonds Fine-Tunes Thrombus Formation

ABSTRACT: Mapping of disulfide bonds and quantification of their redox state in the human histidine rich glycoprotein

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Cell, Blood Plasma, Blood

DISEASE(S): Thrombosis

SUBMITTER: Joyce Chiu

LAB HEAD: Chao Fang

PROVIDER: PXD050718 | Pride | 2024-04-11

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	HRG_disulfide_mapping.mgf	Mgf
	HRG_disulfide_mapping.raw	Raw
	HRG_disulfide_mapping.xlsx	Xlsx
	HRG_only.mgf	Mgf
	HRG_only.raw	Raw

Items per page:

1 - 5 of 13

Similar Datasets

Mechano-redox control of Mac-1 de-adhesion from ICAM-1 by protein disulfide isomerase promotes directional movement under flow

Project description:Quantification of the redox state of disulfide bonds in the beta 2 subunit of human recombinant Mac-1 integrin.

2023-04-09 | PXD032688 | Pride

Alternate covalent form of platelet αIIbβ3

Project description:The αIIbβ3 integrin receptor coordinates platelet adhesion, activation and mechanosensing in thrombosis and haemostasis. Using differential cysteine alkylation and mass spectrometry, we have identified a disulfide bond in the αIIb subunit linking cysteines 490 and 545 that is missing in about one in three integrin molecules on the resting and activated human platelet surface. This alternate covalent form of αIIbβ3 is pre-determined as it is also produced by human megakaryoblasts and bovine hamster kidney cells transfected with recombinant human integrin. From co-immunoprecipitation experiments, the alternate form selectively partitions into focal adhesions on the activated platelet surface. Its function was evaluated in bovine hamster kidney cells expressing a mutant integrin with an ablated C490-C545 disulfide bond. The disulfide mutant integrin has extended residency time in focal adhesions due to reduced rate of clathrin-mediated integrin internalisation and recycling that is associated with enhanced affinity of the αIIb subunit for clathrin adaptor protein-2. The alternate covalent form also has different conformational dynamics measured by monoclonal antibody binding and molecular dynamic simulations, which is reflected in reduced fibrinogen binding and outside-in signalling. These findings indicate that the αIIbβ3 integrin receptor is produced in different covalent forms that have different functions. The C490, C545 cysteine pair is conserved across all 18 integrin α subunits and the disulfide bond in αV in a cancer cell line is similarly missing, suggesting that this alternate integrin form and function is also conserved.

2022-02-17 | PXD025907 | Pride

Patient anti-FVIII drug antibodies bind preferentially to a subset of FVIII covalent states

Project description:Haemophilia A is a chronic life-threatening condition caused by deficiency or dysfunction of plasma coagulation factor VIII (FVIII) and prophylaxis by regular infusion of FVIII protein is a common treatment. A major obstacle to FVIII replacement therapy is the generation of alloantibodies that diminish efficacy. Disulfide bonds link pairs of cysteine residues in proteins and in several proteins have been found to be only partially formed in the mature proteins. FVIII contains 8 disulfide bonds and their redox state in human blood and recombinant FVIII was determined using differential cysteine alkylation and mass spectrometry. All 8 disulphide bonds were found to be unformed in 1 in 10 to 2 in 3 molecules of FVIII populations, which suggested a conformational flexibility that could favour binding of certain ligands to subsets of FVIII with more or less formed disulfide bonds. To test this hypothesis, the binding of a panel of five patient-derived anti-FVIII antibodies to the population of FVIII disulfide-bonded states was evaluated. All five antibodies bound preferentially to FVIII states where 2 or 3 of the 8 disulphides are significantly more unformed; C1918-C1922 in the A3 domain, C2040-C2188 in the C1 domain and C2193-C2345 in the C2 domain. Disulfide bond mutagenesis experiments and molecular dynamics simulations indicate that this subset of FVIII states have long-range conformational dynamism that favours anti-drug antibody binding. These findings will assist efforts to engineer a FVIII molecule that diminishes the emergence of inhibitors and has general implications for autoimmune conditions and antibody drug efficacy.

2025-08-04 | PXD060297 | Pride

In-cell isotope-coded labeling and LC-MS/MS analysis probe warfarin inhibition mechanism

Project description:Warfarin targets human vitamin K epoxide reductase (hVKOR), a redox enzyme in the membrane of endoplasmic reticulum (ER), to prevent the formation of blood clots. Although warfarin has been a popular medication since 1954, its mechanism of action is still unclear. A fundamental issue is the controversial orientation of transmembrane helices (TM) in hVKOR. Stable isotope-coded reagents were usedto label VKOR in free, mutated, and warfarin-binding states directly in the cellular environment, followed by LC-MS/MS bottom-up approach to investigate the warfarin binding mechanism.

2017-03-30 | PXD003774 | Pride

Deamidation drives molecular aging of the SARS-CoV-2 spike receptor-binding motif

Project description:The spike is the main protein component of the SARS-CoV-2 virion surface. The spike receptor-binding motif mediates recognition of the hACE2 receptor, a critical infection step, and is the preferential target for spike-neutralizing antibodies. Post-translational modifications of the spike receptor-binding motif can modulate viral infectivity and immune response. We studied the spike protein searching for asparagine deamidation, a spontaneous event that leads to the appearance of aspartic and isoaspartic residues, affecting both the protein backbone and its charge. We used computational prediction and biochemical experiments to identify five deamidation hotspots in the SARS-CoV-2 spike. Asparagine residues 481 and 501 from the receptor-binding motif deamidate with a half-time of 16.5 and 123 days at 37 °C, respectively. Deamidation is significantly slowed down at 4 °C, pointing at a strong dependence of spike molecular aging on the environmental conditions. We demonstrate that topological constraints drive the conservation of deamidation hotspots among Sarbecoviruses spike proteins. Deamidation of the spike receptor-binding motif decreases the equilibrium constant for binding to the hACE2 receptor more than 3.5-fold, yet its high conservation pattern suggests some positive effect on viral fitness. We propose a model for deamidation of the full SARS-CoV-2 virion that illustrates how deamidation of the spike receptor-binding motif leads to the accumulation on the virion surface of a non-negligible chemically diverse spike population in a timescale of days. Our findings provide a mechanism for molecular aging of the spike, with significant consequences for understanding virus infectivity and vaccine development.

2022-02-17 | PXD027873 | Pride

Deamidation drives molecular aging of the SARS-CoV-2 spike receptor-binding motif

2022-02-15 | PXD028071 | Pride

Semi-Synthesis of an Evasin from Tick Saliva Reveals a Critical Role of Tyrosine Sulfation for Chemokine Binding and Inhibition

Project description:Blood feeding arthropods produce anti-inflammatory salivary proteins called evasins that function through inhibition of chemokine-receptor signalling in the host. Herein, we show that the evasin ACA-01 from the Amblyomma cajennense tick can be post-translationally sulfated at two tyrosine residues, albeit as a mixture of sulfated variants. Homogenously sulfated variants of the proteins were efficiently assembled via a semi-synthetic native chemical ligation strategy. Sulfation significantly improved the binding affinity of ACA-01 for a range of pro-inflammatory chemokines and also enhanced the ability of ACA-01 to inhibit chemokine signalling through cognate receptors. Comparisons of evasin sequences and structural data suggest that tyrosine sulfation serves as a receptor mimetic strategy for recognizing and suppressing the pro-inflammatory activity of a wide variety of mammalian chemokines. As such, the use incorporation of this PTM or mimics thereof into evasins may provide a strategy to optimize these salivary tick proteins for anti-inflammatory applications

2020-05-15 | PXD016778 | Pride

Subtleties in Clathrin Heavy Chain Binding Boxes provide selectivity among Adaptor Proteins of Budding Yeast

Project description:Clathrin, forming the triskelion network, orchestrates highly regulated cellular processes facilitating cargo internalization and trafficking in eukaryotes, with its N-terminal domain (NTD) pivotal for adaptor protein (AP) interactions. The NTD contains up to four AP-binding sites, and their roles in preferential occupancy by APs have not been addressed. Here we present Native MS data showing competition between peptides from Adaptor proteins Ent1 and Ent2.

2024-11-05 | PXD052864 | Pride

LC-MS targeting the rpL40-Lys22 spanning peptide in rpL40-immunoprecipitated representative protein lysates from tissue biopsies of normal stomach and gastric adenocarcinoma (GAC) from KPPC GAC model mice

Project description:We identified SMYD5 (SET and MYND Domain-Containing Protein 5) physiologically methylates core ribosomal protein L40 at lysine 22 (rpL40K22me3).In mouse GAC model, we observed rpL40K22me3 stoichiometry is saturating in both normal and GAC tissue.

2024-06-04 | PXD052359 | Pride

LC-MS targeting the rpL40-Lys22 spanning peptide in rpL40-immunoprecipitated KKLS human gastric cancer cell extracts

Project description:We identified SMYD5 (SET and MYND Domain-Containing Protein 5) physiologically methylates core ribosomal protein L40 at lysine 22 (rpL40K22me3). rpL40K22me3 depletion upon SMYD5 knockdown in GAC KKLS cells was observed by targeted-mass spectrometry of immunoprecipitated rpL40.

2024-06-04 | PXD052358 | Pride