Dataset Information

Developing Chemical Tools for a Systematic Exploration of the Ubiquitin-like Protein FUBI

ABSTRACT: Ubiquitin-like proteins constitute a diverse group of post-translational modifiers that share structural and functional similarities with Ubiquitin, but play distinct roles in cellular signaling. The ubiquitin-like protein FUBI acts as a positive regulator of apoptosis in cancers, where its down-regulation is significantly associated with poor prognosis. Despite the clinical and biological relevance of FUBI-related functions, the intricate regulatory mechanisms orchestrated by FUBI remain elusive. To address this knowledge gap, we developed a linear synthetic platform to generate FUBI-based chemical tools. This platform enables the site-specific incorporation of unnatural building blocks and the introduction of fluorophores, tags, reactive warheads through N- and C-terminal labeling strategies. Using this platform, we created activity-based probes (ABPs) for FUBI conjugation and deconjugation enzymes, validating them in cell lysate-based assays and proteomics. Additionally, we synthesized K25 triazole-linked Di-FUBI as bioisosteric uncleavable mimic of native Di-FUBI to study the proteolytic cleavage of FUBI chains. Collectively, these ABPs demonstrate the versatility and specificity of our synthetic FUBI platform, advancing the characterization of FUBI-related enzymes and decoding their roles in cellular processes at the molecular level.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Rayman Tjokrodirijo

LAB HEAD: Peter A. van Veelen

PROVIDER: PXD057422 | Pride | 2025-06-25

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

Items per page:

1 - 5 of 183

Similar Datasets

Project description:Comparative, dose-dependent analysis of interactions between small molecule drugs and their targets, as well as off-targets, in complex proteomes is crucial for selecting optimal drug candidates. The affinity of small molecules for targeted proteins is largely dictated by interactions between amino acid side chains and these drugs. Thus, studying drug-protein interactions at an amino acid resolution provides a comprehensive understanding of drug selectivity and efficacy. In this study, we further refined the site-specific activity-based protein profiling strategy, PhosID-ABPP, on a timsTOF Pro mass spectrometer. This refinement enables dose-dependent competition of inhibitors within a single cellular proteome. Here, a comparative analysis of two activity-based probes (ABPs), developed to selectively target the epidermal growth factor receptor (EGFR), namely PF-06672131 and PF-6422899, facilitated the simultaneous identification of ABP-specific binding sites at a proteome-wide scale within a cellular proteome. Dose-dependent probe-binding preferences for proteinaceous cysteines, even at low nanomolar ABP concentrations, could be revealed. Notably, while both ABPs showed comparable affinities for the EGFR, PF-06672131 had a broader off-target reactivity profile. In contrast, PF-6422899 exhibited higher affinity for the ERBB2 receptor and bound to catalytic cysteines in several other enzymes, which is likely to disrupt their catalytic activity. Notably, PF-06672131 also effectively labeled ADP/ATP translocase proteins at a concentration of just 1 nanomolar. Additionally, analysis of different binding sites within the EGF receptor and the voltage-dependent anion channel 2 revealed secondary binding sites of both probes and provided insights into the binding poses of inhibitors on these proteins. Insights from the PhosID-ABPP analysis of these two ABPs serve as a valuable resource for understanding drug on- and off-target engagement in a dose- and site-specific manner.

			Action	DRS
	E20232003235.raw	Raw
	E20232003236a.raw	Raw
	E20232003237a.raw	Raw
	E20232003238a.raw	Raw
	E20232003239a.raw	Raw

Dataset Information

Developing Chemical Tools for a Systematic Exploration of the Ubiquitin-like Protein FUBI

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets