Chemical Proteomic Discovery of Isotype‐Selective Covalent Inhibitors of the RNA Methyltransferase NSUN2

Angewandte Chemie International Edition, Journal Year: 2023, Volume and Issue: 62(51)

Published: Nov. 1, 2023

Abstract 5‐Methylcytosine (m 5 C) is an RNA modification prevalent on tRNAs, where it can protect tRNAs from endonucleolytic cleavage to maintain protein synthesis. The NSUN family (NSUN1‐7 in humans) of methyltransferases are capable installing the methyl group onto C position cytosines RNA. NSUNs implicated a wide range (patho)physiological processes, but selective and cell‐active inhibitors these enzymes lacking. Here, we use cysteine‐directed activity‐based profiling (ABPP) discover azetidine acrylamides that act as stereoselective covalent human NSUN2. Despite targeting conserved catalytic cysteine family, NSUN2 show negligible cross‐reactivity with other exhibit good proteome‐wide selectivity. We verify inhibit activity recombinant NSUN2, not NSUN6, demonstrate compounds stereoselectively disrupt NSUN2‐tRNA interactions cancer cells, leading global reduction tRNA m content. Our findings thus highlight potential create isotype‐selective chemistry cysteine.

Language: Английский

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Redirecting the pioneering function of FOXA1 with covalent small molecules

Molecular Cell, Journal Year: 2024, Volume and Issue: unknown

Published: Oct. 1, 2024

Language: Английский

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Proteomic Ligandability Maps of Phosphorus(V) Stereoprobes Iden-tify Covalent TLCD1 Inhibitors

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 31, 2025

ABSTRACT Activity-based protein profiling (ABPP) of stereoisomerically defined sets electrophilic compounds (‘stereoprobes’) offers a versatile way to discover covalent ligands for proteins in native biological systems. Here we report the synthesis and chemical proteomic characterization stereoprobes bearing P(V)-oxathiaphospholane (OTP) reactive group. ABPP experiments identified numerous human cancer cells that showed stereoselective reactivity with OTP stereoprobes, confirmed several these liganding events recombinant proteins. engaging poorly characterized transmembrane TLCD1 impaired incorporation monounsaturated fatty acids into phosphatidylethanolamine lipids cells, lipidomic phenotype mirrored genetic disruption this protein. Using AlphaFold2, found structurally resembles ceramide synthase acid elongase families coenzyme A-dependent lipid processing enzymes. This structural similarity included conservation catalytic histidine residues, mutation which blocked stereoprobe remodeling activity TLCD1. Taken together, data indicate acts as acyltransferase function inhibitors enzymatic activity. Our findings thus illuminate how analysis can facilitate functional annotation inhibition key metabolic enzyme cells.

Language: Английский

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Ligand discovery by activity-based protein profiling

Cell chemical biology, Journal Year: 2024, Volume and Issue: 31(9), P. 1636 - 1651

Published: Sept. 1, 2024

Language: Английский

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Chemical tools to expand the ligandable proteome: Diversity-oriented synthesis-based photoreactive stereoprobes

Cell chemical biology, Journal Year: 2024, Volume and Issue: unknown

Published: Nov. 1, 2024

Language: Английский

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Protein Electrostatic Properties are Fine-Tuned Through Evolution

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: April 21, 2025

Abstract Protein ionization states provide electrostatic forces to modulate protein structure, stability, solubility, and function. Until now, predicting understanding electrostatics have relied on structural information. Here we demonstrate that primary sequence alone enables remarkably accurate p K a predictions through KaML-ESM, model leverages evolutionary representations from ultra-large language models ESMs pretraining with synthetic dataset. The KaML-ESM achieves RMSEs approaching the experimental precision limit of ∼0.5 pH units for Asp, Glu, His, Lys residues, while reducing Cys prediction errors 1.1 – further improvement expected as training dataset expands. state-of-the-art performance was validated external evaluations, including proteome-wide analysis values. Our results support notation encodes not only structure function but also properties, which may been co-optimized evolution. Lastly, KaML, sequence-based end-to-end ML platform researchers map landscapes, facilitating applications ranging drug design engineering molecular simulations.

Language: Английский

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Proteomic Ligandability Maps of Phosphorus(V) Stereoprobes Identify Covalent TLCD1 Inhibitors

Journal of the American Chemical Society, Journal Year: 2025, Volume and Issue: unknown

Published: April 23, 2025

Activity-based protein profiling (ABPP) of stereoisomerically defined sets electrophilic compounds ('stereoprobes') offers a versatile way to discover covalent ligands for proteins in native biological systems. Here we report the synthesis and chemical proteomic characterization stereoprobes bearing P(V)-oxathiaphospholane (OTP) reactive group. ABPP experiments identified numerous human cancer cells that showed stereoselective reactivity with OTP stereoprobes, confirmed several these liganding events recombinant proteins. engaging poorly characterized transmembrane TLCD1 impaired incorporation monounsaturated fatty acids into phosphatidylethanolamine lipids cells, lipidomic phenotype mirrored genetic disruption this protein. Using AlphaFold2, found structurally resembles ceramide synthase acid elongase families coenzyme A-dependent lipid processing enzymes. This structural similarity included conservation catalytic histidine residues, mutation which blocked stereoprobe remodeling activity TLCD1. Taken together, data indicate acts as acyltransferase function inhibitors enzymatic activity. Our findings thus illuminate how analysis can facilitate functional annotation inhibition key metabolic enzyme cells.

Language: Английский

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Towards Site-specific Manipulation in Cysteine-mediated Redox Signaling

Chemical Science, Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 1, 2025

This perspective summarized emerging chemical biology strategies to interrogate cysteine-mediated redox signaling in a site-specific fashion.

Language: Английский

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Advancing Covalent Ligand and Drug Discovery beyond Cysteine

Chemical Reviews, Journal Year: 2025, Volume and Issue: unknown

Published: May 22, 2025

Targeting intractable proteins remains a key challenge in drug discovery, as these often lack well-defined binding pockets or possess shallow surfaces not readily addressed by traditional design. Covalent chemistry has emerged powerful solution for accessing protein sites difficult to ligand regions. By leveraging activity-based profiling (ABPP) and LC-MS/MS technologies, academic groups industry have identified cysteine-reactive ligands that enable selective targeting of challenging modulate previously inaccessible biological pathways. Cysteines within are rare, however, developing covalent target additional residues hold great promise further expanding the ligandable proteome. This review highlights recent advancements amino acids beyond cysteine with an emphasis on tyrosine- lysine-directed their applications chemical biology therapeutic development. We outline process using proteomic methodology, highlighting successful examples discuss considerations future expansion acid proteins.

Language: Английский

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An allosteric cyclin E-CDK2 site mapped by paralog hopping with covalent probes

Nature Chemical Biology, Journal Year: 2024, Volume and Issue: unknown

Published: Sept. 18, 2024

Language: Английский

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