title: A Cell-Permeable Biscyclooctyne As a Novel Probe for the Identification of Protein Sulfenic Acids.
abstract: Reactive oxygen species act as important second messengers in cell signaling and homeostasis through the oxidation of protein thiols. However, the dynamic nature of protein oxidation and the lack of sensitivity of existing molecular probes have hindered our understanding of such reactions; therefore, new tools are required to address these challenges. We designed a bifunctional variant of the strained bicyclo[6.1.0]nonyne (BCN-E-BCN) that enables the tagging of intracellular protein sulfenic acids for biorthogonal copper-free click chemistry. In validation studies, BCN-E-BCN binds the sulfenylated form of the actin-severing protein cofilin, while mutation of the cognate cysteine residues abrogates its binding. BCN-E-BCN is cell permeable and reacts rapidly with cysteine sulfenic acids in cultured cells. Using different azide-tagged conjugates, we demonstrate that BCN-E-BCN can be used in various applications for the detection of sulfenylated proteins. Remarkably, cycloaddition of an azide-tagged fluorophore to BCN-E-BCN labeled proteins produced in vivo can be visualized by fluorescence microscopy to reveal their localization. These findings demonstrate a novel and multifaceted approach to the detection and trapping of sulfenic acids.
authors: Cancer Research UK Beatson Institute , Garscube Estate, Switchback Road, Glasgow G61 1BD, United Kingdom.
authors: David J
authors: DJ
authors: McGarry
authors: WestCHEM School of Chemistry, University of Glasgow , Glasgow G12 8QQ, United Kingdom.
authors: Maria M
authors: MM
authors: Shchepinova
authors: Cancer Research UK Beatson Institute , Garscube Estate, Switchback Road, Glasgow G61 1BD, United Kingdom.
authors: Sergio
authors: S
authors: Lilla
authors: WestCHEM School of Chemistry, University of Glasgow , Glasgow G12 8QQ, United Kingdom.
authors: Richard C
authors: RC
authors: Hartley
authors: Cancer Research UK Beatson Institute , Garscube Estate, Switchback Road, Glasgow G61 1BD, United Kingdom.|Institute of Cancer Sciences, University of Glasgow , Glasgow G12 8QQ, United Kingdom.
authors: Michael F
authors: MF
authors: Olson
mesh_terms: D051339:Actin Depolymerizing Factors / Q000737:chemistry; D001386:Azides / Q000737:chemistry*; D015153:Blotting, Western; D001643:Bridged Bicyclo Compounds / Q000737:chemistry*; D045744:Cell Line, Tumor; D057930:Click Chemistry; D006801:Humans; D007202:Indicators and Reagents / Q000737:chemistry; D008856:Microscopy, Fluorescence; D015335:Molecular Probes / Q000737:chemistry; D011506:Proteins / Q000737:chemistry*; D013434:Sulfenic Acids / Q000032:analysis*
chemical_list: D051339:Actin Depolymerizing Factors; D001386:Azides; D001643:Bridged Bicyclo Compounds; D007202:Indicators and Reagents; D015335:Molecular Probes; D011506:Proteins; D013434:Sulfenic Acids; C556617:bicyclo(6.1.0)nonyne