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Proteomic discovery of chemical probes that perturb protein complexes in human cells II

ABSTRACT: Most proteins in the human proteome lack chemical probes, and several large-scale and generalizable small-molecule binding assays have been introduced to address this problem. How compounds discovered in such “binding-first” assays affect protein function, nonetheless, often remains unclear. Here, we describe a “function first” proteomic strategy that uses size exclusion chromatography (SEC) to assess the global impact of electrophilic compounds on protein complexes in human cells. Integrating the SEC data with cysteine-directed activity-based profiling identifies changes in protein-protein interactions that are caused by site-specific liganding events, including the stereoselective engagement of cysteines in PSME1 and SF3B1 that disassemble the PA28 proteasome regulatory complex and remodel the spliceosome, respectively. Our findings thus show how multidimensional proteomic analysis of focused libraries of electrophilic compounds can expedite the discovery of chemical probes with site-specific functional effects on protein complexes in human cells. In this experiment, we probed the RNA-binding profile of DDX42 by eCLIP via an endogenously introduced HA-tag in the context of various SF3B1 ligands.

ORGANISM(S): Homo sapiens

PROVIDER: GSE220845 | GEO | 2023/04/13

REPOSITORIES: GEO

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Proteomic discovery of chemical probes that perturb protein complexes in human cells

Project description:Most proteins in the human proteome lack chemical probes, and several large-scale and generalizable small-molecule binding assays have been introduced to address this problem. How compounds discovered in such “binding-first” assays affect protein function, nonetheless, often remains unclear. Here, we describe a “function first” proteomic strategy that uses size exclusion chromatography (SEC) to assess the global impact of electrophilic compounds on protein complexes in human cells. Integrating the SEC data with cysteine-directed activity-based profiling identifies changes in protein-protein interactions that are caused by site-specific liganding events, including the stereoselective engagement of cysteines in PSME1 and SF3B1 that disassemble the PA28 proteasome regulatory complex and remodel the spliceosome, respectively. Our findings thus show how multidimensional proteomic analysis of focused libraries of electrophilic compounds can expedite the discovery of chemical probes with site-specific functional effects on protein complexes in human cells. In this experiment, we probed the RNA-binding profile of DDX42 by eCLIP via an endogenously introduced HA-tag in the context of various SF3B1 ligands.

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Proteomic discovery of chemical probes that perturb protein complexes in human cells

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Proteomic discovery of chemical probes that perturb protein complexes in human cells

Project description:Proteomic discovery of chemical probes that perturb protein complexes in human cells

| PRJNA909188 | ENA

Proteomic discovery of chemical probes that perturb protein complexes in human cells II

Project description:Proteomic discovery of chemical probes that perturb protein complexes in human cells II

| PRJNA911743 | ENA

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