Copper(I)-Nitrene Platform for Chemoproteomics Profiling of Methionine

ABSTRACT: Methionine plays a critical role in various biological and cell regulatory processes, making its chemoproteomic profiling indispensable for exploring its functions and potential in protein therapeutics. However, the advancement of methionine chemoproteomics is currently hindered by the scarcity of efficient labeling strategies. Methionine exhibits high kinetic reactivity and undergoes rapid oxidation due to the lower energy gap between Lowest Unoccupied Molecular Orbital (LUMO) and Highest Occupied Molecular Orbital (HOMO). Building on this unique high reactivity of methionine (Met), we introduce Copper(I)-Nitrene Platform (CuNiP) for robust, and selective labeling of Met to generate highly stable sulfonyl sulfimide conjugates under physiological conditions. We demonstrated the versatile proficiency of CuNiP to label methionine in a wide range of bioactive peptides, intact proteins (6.5-79.5 kDa) independent of molecular weight and 3D structures and proteins in complex cell lysate mixtures with varying payloads. Leveraging nitrene reactivity-based probes, we have discovered an array of new, previously undiscovered, ligandable proteins and sites harboring hyperreactive methionine within the landscape of the human proteome. Crucially, we employed CuNiP for live cell chemoproteomic profiling of methionine in breast (T47D) and prostate cancer (LnCap) cell lines, observing minimal cytotoxic effects and achieving dose-dependent labeling. Confocal imaging further revealed the spatial distribution of modified proteins within cell membrane, cytoplasm, and nucleus, underscoring the platform's potential in profiling the cellular interactome. This breakthrough, with its insights into live cell methionine labeling will deepen our understanding of protein biochemistry and also pioneers a transformative pathway in drug discovery.

INSTRUMENT(S): Orbitrap Fusion Lumos, Bruker Daltonics timsTOF series

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: David Gordon  

LAB HEAD: David Ezra Gordon

PROVIDER: PXD051224 | Pride | 2024-04-24

REPOSITORIES: Pride