Proteomics

Dataset Information

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Chemoproteomic Profiling by Cysteine Fluoroalkylation Reveals the DNA Repair Protein XRCC5 as a Functional Target of Myrocin G


ABSTRACT: Chemoproteomic profiling of cysteines has emerged as a powerful method for screening the proteome-wide targets of cysteine-reactive frag-ments, drugs and natural products. Herein, we report the development and an in-depth evaluation of a tetrafluoroalkyl benziodoxole as a cyste-ine-selective chemoproteomic probe. We show that this probe features numerous key improvements compared to the traditionally used cyste-ine-reactive probes, including a superior target occupancy, faster labeling kinetics, and broader proteomic coverage thus enabling profiling of cysteines directly in live cells. Further, the fluorine ‘signature’ of probe 7 constitutes an additional advantage resulting in a more confident adduct-amino acid site assignment in mass spectrometry-based identification workflows. We demonstrate the utility of our new probe for proteome-wide target profiling by identifying the cellular targets of (−)-myrocin G, an antiproliferative fungal natural product with a to-date unknown mechanism of action. We show that this natural product and a simplified analog target the X-ray repair cross-complementing protein 5 (XRCC5), an ATP-dependent DNA helicase that primes DNA repair machinery for non-homologous end joining (NHEJ) upon DNA double strand breaks, making them the first reported inhibitors of this biomedically highly important protein. We further demonstrate that myrocins disrupt the interaction of XRCC5 with DNA leading to sensitization of cancer cells to the chemotherapeutic agent etoposide as well as UV-light induced DNA damage. Altogether, our next generation cysteine-reactive probe enables broader and deeper profiling of the cyste-inome rendering it a highly attractive tool for elucidation of targets of electrophilic small molecules.

INSTRUMENT(S): Q Exactive HF-X

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture

DISEASE(S): Cervix Carcinoma

SUBMITTER: Daniel Abegg  

LAB HEAD: Alexander Adibekian

PROVIDER: PXD029255 | Pride | 2021-12-13

REPOSITORIES: Pride

Dataset's files

Source:
Action DRS
180129_HeLa_2mM-TogniC2F5_50mM-IAA.raw Raw
180129_HeLa_DMSO_50mM-IAA.raw Raw
180306_HeLa_2mM-TogniC2F5_50mM-IAA.msf Msf
180306_HeLa_2mM-TogniC2F5_50mM-IAA.raw Raw
180306_HeLa_DMSO_50mM-IAA.raw Raw
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Publications

Chemoproteomic Profiling by Cysteine Fluoroalkylation Reveals Myrocin G as an Inhibitor of the Nonhomologous End Joining DNA Repair Pathway.

Abegg Daniel D   Tomanik Martin M   Qiu Nan N   Pechalrieu Dany D   Shuster Anton A   Commare Bruno B   Togni Antonio A   Herzon Seth B SB   Adibekian Alexander A  

Journal of the American Chemical Society 20211124 48


Chemoproteomic profiling of cysteines has emerged as a powerful method for screening the proteome-wide targets of cysteine-reactive fragments, drugs, and natural products. Herein, we report the development and an in-depth evaluation of a tetrafluoroalkyl benziodoxole (TFBX) as a cysteine-selective chemoproteomic probe. We show that this probe features numerous key improvements compared to the traditionally used cysteine-reactive probes, including a superior target occupancy, faster labeling kine  ...[more]

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