Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Harnessing Ionic Selectivity In Acetyltransferase Chemoproteomic Probes

ABSTRACT: Chemical proteomics (chemoproteomics) has emerged as a powerful strategy for the high-throughput annotation of protein function. This approach involves using an active site probe to capture an enzyme class of interest from cells, allowing the biological activity of multiple enzymes within that family to be assessed in parallel. Coenzyme A based chemoproteomic probes may have utility in studying the pharmacological interactions of additional enzyme classes, beyond histone acetyltransferases. However, one challenge in applying this approach is how to differentiate a ‘true hit’ from background. Many histone acetyltransferases utilize an ordered binding mechanism, with acetyl-CoA binding first, which allows their chemoproteomic capture to be competed by pre-incubating lysates with acetyl-CoA. However, for acetyltransferases that don’t exhibit this binding mode, or which bind to a different ligand, acetyl-CoA competition may not be evident, limiting our ability to discern selective chemoproteomic enrichment from noise. In this study, two physiochemically distinct capture probes were used to enrich proteins to distinguish true hits from background and identify new targets selectively enriched by CoA-based probes, thus expanding the scope of protein-ligand interactions open to chemoproteomic interrogation.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Bone Marrow Cell

SUBMITTER: Michaella Levy

LAB HEAD: Laurence Florens

PROVIDER: PXD020856 | Pride | 2021-08-19

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	Hs_Ahx-4plex_K562-WCE_KAT-1_Ti_QE1_1.raw	Raw
	Hs_Ahx-4plex_K562-WCE_KAT-2_Ti_QE1_1.raw	Raw
	Hs_Ahx-4plex_K562-WCE_KAT-3_Ti_QE1_1.raw	Raw
	Hs_Ahx-4plex_K562-WCE_KAT_QE1_6mods_NAb_Kac_2.mgf	Mgf
	Hs_Ahx-4plex_K562-WCE_KAT_QE1_6mods_NAb_Kac_2.mzML	Mzml

Items per page:

1 - 5 of 8

Similar Datasets

Investigating peptide-Coenzyme A-conjugates as chemical probes for proteomic profiling of N-terminal and lysine acetyltransferases

Project description:Acetyl groups are transferred from Acetyl-Coenzyme A (Ac-CoA) to protein N-termini and lysine side chains by N-terminal acetyltransferases (NATs) and lysine acetyltransferases (KATs), respectively. Building on lysine-CoA conjugates as KAT probes, we have synthesized peptide probes with CoA conjugated to N-terminal alanine (α-Ala-CoA), proline (α-Pro-CoA) or tri-glutamic acid (α-3Glu-CoA) units for interactome profiling of NAT complexes. These probes recruited catalytic and auxiliary subunits of the major NAT complexes from cell lysates specifically. The α-Ala-CoA probe was more specific in recruiting NAT catalytic and auxiliary subunits when compared to a lysine CoA-conjugate which bound only a subset of endogenous KATs. Interactome profiling with the α-Pro-CoA probe showed reduced NAT recruitment in favor of metabolic CoA binding proteins, while α-3Glu-CoA steered the interactome towards NAA80 and NatB. These findings agreed with the inherent substrate specificities of the target proteins and showed that N-terminal CoA-conjugated peptides are versatile probes for NAT complex profiling in lysates of physiological and pathological backgrounds.

2022-05-02 | PXD032957 | JPOST Repository

Chemical proteomics profiling of N-terminal and lysine acetyltransferases

Project description:Acetylation of amino groups is a prevalent protein modification in all kingdoms of life. Acetyl groups are transferred from Coenzyme A (CoA) to protein N-termini and lysine side chains by N-terminal acetyltransferases (NATs) and lysine acetyltransferases (KATs), respectively. Building on lysine-CoA conjugates as KAT probes, we have synthesized N-terminal CoA-conjugated peptide probes for interactome profiling of NAT complexes. These probes specifically recruited catalytic and auxiliary subunits of the major NAT complexes from cell lysates. When comparing the specificity of N-terminal and lysine CoA-conjugated probes the latter bound only a subset of endogenous KATs and appeared more prone to recruitment of other CoA-binding proteins. Western blot validation confirmed the specificity of selected NATs and KATs towards these probes supporting the notion that N-terminal CoA-conjugated peptides are versatile probes for NAT complex profiling in lysates of physiological and pathological backgrounds.

2021-04-14 | PXD018544 | JPOST Repository

Characterization of lysine acetylation in Toxoplasma gondii proteome lacking acetyl-CoA transporter 1 (AT1).

Project description:Acetyl-CoA participates in post-translational modification of proteins, central carbon and lipid metabolism in several cell compartments. In mammals, the acetyl-CoA transporter 1 (AT1) facilitates the flux of cytosolic acetyl-CoA into the endoplasmic reticulum (ER), enabling the acetylation of proteins of the secretory pathway, in concert with dedicated acetyltransferases including Nat8. However, the implication of the ER acetyl-CoA pool in acetylation of ER-transiting proteins and their relevance throughout the parasites’ life cycle is unknown. Here, we evaluated the impact of blocking putative cytosolic acetyl-CoA import on acetylation of proteins through KO of the homologue of AT1 in the parasite Toxoplasma gondii.

2022-07-25 | PXD030932 | Pride

Nucleoside-Diphosphate Kinase 1 and 2 are master regulators of a liver protective response to high fat diet [ChIP-seq]

Project description:Fatty acid homeostasis is critical for normal cellular physiology and leads to severe diseases when deregulated. Here we report Nucleoside-Diphosphate Kinase 1 and 2 (NME1/2) as major cellular co-enzyme A (CoA) and acetyl-CoA binding proteins and negative regulators of de novo lipogenesis (DNL). Structural studies demonstrate that Nme1 recognizes CoA through its nucleotide moiety, which competes for binding with ADP/ATP. By using a Nme2 ko mouse model, we observe that NME2 is required for the gene transcriptional response to a high fat diet (HFD) in liver cells, leading to a repression of lipogenesis and the activation of a protective gene response. Nme2 ko mice submitted to a HFD challenge are unable to repress key lipogenic genes, resulting in an excessive triglyceride synthesis and liver steatosis. Mechanistically, the NME2-dependent down-regulation of DNL in response to HFD changes the balance for the use of acetyl-CoA between the competing paths of acetylation and DNL, thereby increasing TSS-targeted histone acetylation and gene activation. A structure-guided generation of a NME1 mutant, with intact NDK activity, but unable to bind CoA, directly demonstrates the functional impact of acetyl-CoA/CoA binding by NME1/2 in repressing DNL. Taken together, these findings highlight a yet unknow protective liver response to HFD, regulated by multi-ligand binding proteins which act as direct sensors for the cellular levels of NDP/NTP and CoA/acetyl-CoA.

2023-08-01 | GSE198687 | GEO

Nucleoside-Diphosphate Kinase 1 and 2 are master regulators of a liver protective response to high fat diet [RNA-seq]

2023-08-01 | GSE198686 | GEO

RNA-sequencing analysis of glucose and acetate regulated transcripts in glioblastoma cells

Project description:Our studies indicate that glucose and acetate can regulate histone acetylation by altering the acetyl-CoA concentrations in the cell. The purpose of this study was to to determine whether specific gene sets correlated with acetyl-CoA availability. We conclude that 10% of glucose-regulated genes are acetyl-CoA regulated genes (genes suppressed or induced by low glucose and reversed by acetate). Acetate usually regulated gene expression in the same direction as glucose, suggesting that acetyl-CoA is a key mediator of glucose-dependent gene expression. The experiments were performed in quadruplicates for each condition with a total of 12 samples

2014-08-05 | E-GEOD-57488 | biostudies-arrayexpress

Cytosolic acetyl-CoA promotes histone acetylation predominantly at H3K27 in Arabidopsis

Project description:Acetyl-Coenzyme A (acetyl-CoA) is a central metabolite and the acetyl source for protein acetylation, particularly histone acetylation that promotes gene expression. However, the effect of acetyl-CoA levels on histone acetylation status in plants remains unknown. Here, we show that malfunctioned cytosolic acetyl-CoA carboxylase1 (ACC1) in Arabidopsis leads to elevated levels of acetyl-CoA and promotes histone hyperacetylation predominantly at lysine 27 of histone H3 (H3K27). The increase of H3K27 acetylation (H3K27ac) is dependent on ATP-citrate lyase which cleaves citrate to acetyl-CoA in the cytoplasm, and requires histone acetyltransferase GCN5. A comprehensive analysis of the transcriptome and metabolome in combination with the genome-wide H3K27ac profiles of acc1 mutants, demonstrate the dynamic changes of H3K27ac, gene transcripts and metabolites occurring in the cell by the increased levels of acetyl-CoA. This study suggests that H3K27ac is an important link between cytosolic acetyl-CoA level and gene expression in response to the dynamic metabolic environments in plants.

2017-09-25 | GSE79524 | GEO

The state transition acetyltransferase GNAT2 interacts with GNAT1 and GNAT3 in independent complexes

Project description:The acetylation of amino groups within proteins is one of the major posttranslational modifications (PTMs) in all organisms. N-terminal acetyltransferases (NATs) or lysine acetyltransferases (KATs) catalyze the transfer of the acetyl group from the donor substrate acetyl coenzyme A (Ac-CoA) to the targeted amino group, which is either the α-amino groups of N-termini or the ε-amino group of internal lysine residues. In contrast to N-terminal acetylation (NTA), the acetylation of lysine residues (KA) is reversible, whereby lysine deacetylases (KDACs) are responsible for the removal of acetyl groups from proteins. Furthermore, KA occurs posttranslationally, whereas NTA is described mainly as a cotranslational modification with the exception for N-terminally processed proteins, which are imported to chloroplasts and mitochondria

2024-08-19 | PXD044167 | JPOST Repository

N-Acetylation of secreted proteins is widespread in Apicomplexa and independent of acetyl-CoA transporter AT1

Project description:Acetyl-CoA critically participates in post-translational modification of proteins, central carbon and lipid metabolism in several compartments of eukaryotic cells. In mammals, the acetyl-CoA transporter 1 (AT1) facilitates the flux of cytosolic acetyl-CoA into the endoplasmic reticulum (ER) enabling the acetylation of proteins of the secretory pathway in concert with dedicated acetyltransferases including Nat8. Homologues of AT1 and Nat8 were identified in the apicomplexan parasites Toxoplasma gondii and Plasmodium berghei. However, the implication of acetyl-CoA pool influx into the ER in acetylation of ER-transiting proteins and their relevance throughout the parasites’ life cycle is unknown. Here, we report proteome-wide analyses, which revealed unprecedented widespread N-terminal acetylation marks of secreted proteins in both parasites. Deletion of the gene coding for AT1 in both parasites, resulted in global loss of fitness and in addition to retardation in erythrocytic development, malaria parasites are blocked in transmission to mosquitoes. However, in absence of AT1 proteome wide lysine and N-terminal acetylation modifications remain unaltered. This highlights a role of AT1 in parasite development uncoupled to acetylation capacity, indicative of an unusually active acetylation machinery occurring in the ER of Apicomplexa.

2022-06-20 | PXD029616 | Pride

Acetyl-CoA flux maintains intracellular crosstalk and modulats engagement of the secretory pathway

Project description:Nε-lysine acetylation has emerged as a central mechanism to maintain quality control and protein homeostasis within the Endoplasmic Reticulum (ER) and secretory pathway. The ER acetylation machinery includes AT-1/SLC33A1, a membrane transporter that translocates acetyl-CoA from the cytosol to the ER lumen, and ATase1 and ATase2, two ER membrane-bound acetyltransferases that acetylate ER cargo proteins. Dysfunctional AT-1, as caused by loss-of-function mutations or gene duplication events, results in neurodevelopmental or neurodegenerative disorders. Experiments in our lab have demonstrated that these human diseases can be effectively recapitulated in mouse models. In this thesis, we used two models of dysregulated acetyl-CoA flux: AT-1S113R/+, a model of AT-1 haploinsufficiency, and AT-1 sTg, a model of systemic overexpression of AT-1. First, we examined upstream processes of cytosol-to-ER acetyl-CoA flux by evaluating the cytosolic pool of acetyl-CoA. The aberrant AT-1 models demonstrated distinct metabolic reprogramming of lipid metabolism and mitochondria bioenergetics. Dysregulated acetyl-CoA flux resulted in global changes at the level of the proteome and the acetyl-proteome. Second, we examined the downstream consequences of cytosol-to-ER acetyl-CoA flux. Specifically, we investigated the engagement and functional organization of the secretory pathway and used N-glycoproteomics to determine the quality of secreted proteins. Aberrant AT-1 models demonstrated reorganization of the ER, Golgi, and ERGIC, as well as a delay in glycoproteins clearing the Golgi apparatus. Additionally, AT-1 sTg mice showed a marked delay in protein trafficking to the cell surface. The N-glycoproteome revealed significant alterations, highlighting changes in the quality of the secretome.

2021-09-09 | PXD019770 | Pride

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data