Proteomics

Dataset Information

0

Engineering Af1521 macro domain increases ADP-ribose binding affinity and leads to the identification of more ADP-ribosylated proteins

ABSTRACT: Protein ADP-ribosylation is a covalent, reversible post-translational modification that has important functions in several cellular mechanisms. The identification of modified proteins in cells has proven challenging and, due to the low abundance of protein ADP-ribosylation, has mainly been possible via enrichment methodologies using ADP-ribose binding domains. Here, using random mutagenesis and in vitro selection with an ADP-ribosylated histone peptide, we engineered the archaeal Af1521 macro domain to generate an engineered isoform containing nine mutations that displays significantly increased affinity towards ADP-ribose. Comparison of the wild-type and the engineered Af1521 macro domains using our proteomic ADP-ribosylome enrichment workflow demonstrated an increased identification rate of ADP-ribosylated proteins with the engineered Af1521.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell

SUBMITTER: Kathrin Nowak  

LAB HEAD: Michael O. Hottiger

PROVIDER: PXD016686 | Pride | 2020-10-20

REPOSITORIES: Pride

Json Xml

Dataset's files

Source:
Action DRS
10mg_eAf_1_EThcD.csv Csv
10mg_eAf_1_HCD.csv Csv
10mg_eAf_2_EThcD.csv Csv
10mg_eAf_2_HCD.csv Csv
10mg_eAf_3_EThcD.csv Csv
Items per page:
1 - 5 of 106
altmetric image

Publications

Protein ADP-ribosylation is a reversible post-translational modification that regulates important cellular functions. The identification of modified proteins has proven challenging and has mainly been achieved via enrichment methodologies. Random mutagenesis was used here to develop an engineered Af1521 ADP-ribose binding macro domain protein with 1000-fold increased affinity towards ADP-ribose. The crystal structure reveals that two point mutations K35E and Y145R form a salt bridge within the A  ...[more]

Similar Datasets

Proteomics Gas-phase Fragmentation of ADP-Ribosylated Peptides: Arginine-Specific Side-Chain Losses and their Implication in Database Searches
2021-05-19 | PXD017013 | Pride
Proteomics A study into the ADP-ribosylome of IFN--stimulated THP-1 human macrophage-like cells identifies ARTD8/PARP14 and ARTD9/PARP9 ADP-ribosylation
2020-01-27 | PXD011690 | Pride
Proteomics Proteomic characterization of the heart and skeletal muscle reveals widespread Arginine ADP-ribosylation by the ectopic ADP-ribosyltransferase ARTC1
2018-08-17 | PXD008041 | Pride
Proteomics ADP-ribosylation site analysis
2016-10-04 | PXD004245 | Pride
Proteomics Affinity purification of ADP-ribosylated proteins from A. thaliana plants expressing the Pseudomonas syringae type III effector HopU1
2024-10-22 | PXD056608 | Pride
Proteomics Pseudomonas type III effector AvrRpm1 ADP-ribosylates Arabidopsis thaliana PHOSPHOLIPASE D GAMMA 3 (PLDGAMMA3)
2024-10-22 | PXD056607 | Pride
Proteomics Pseudomonas type III effector AvrRpm1 ADP-ribosylates Arabidopsis thaliana MEMBRANE-ASSOCIATED MANNITOL-INDUCED (MAMI)
2023-09-28 | PXD045549 | Pride
Proteomics Comparison of proteins identified using the Af1521 Macro domain (Nowak et al., https://doi.org/10.1038/s41467-020-18981-w) versus the non-functional (G42E) Macro domain for enrichment of ADP-ribosylated proteins from A. thaliana plants expressing the Pseudomonas syringae type III effectors AvrRpm1 or HopF2 (DDA method)
2023-09-28 | PXD045726 | Pride
Proteomics An advanced strategy for comprehensive profiling of ADP-ribosylation sites using mass spectrometry-based proteomics
2019-02-26 | PXD012243 | Pride
Proteomics Lack of the mitochondrial NAD transporter SLC25A51 redistributes NAD+ compartmentalization and cellular ADP-ribosylation
2023-10-24 | PXD031373 | Pride