Sodium ascorbate

4-vinylbenzyl chloride (Sigma Aldrich, Buchs, Switzerland), DMF (VWR, Schlieren, Switzerland), sodium azide (Sigma Aldrich), diethyl ether (Sigma Aldrich), MgSO₄ (Sigma Aldrich), sodium dodecyl sulfate (Sigma Aldrich), propargyl glicidyl ether (Sigma Aldrich), propargyl-dPEG1-NHS–ester (Rapp Polymer, Tübingen, Germany), sodium ascorbate (Sigma Aldrich), Protein A (Syd Labs, Natick, MA, USA), Cys-Terminated protein A (ProspecBio, East Brunswick, NJ, USA), styrene (Sigma Aldrich), divinyl benzene (Sigma Aldrich), potassium persulfate (Sigma Aldrich), 2,2’-azobis(2-methylpropionitrile) (AIBN) (Sigma Aldrich), copper sulfate (Sigma Aldrich), sodium ascorbate (Sigma Aldrich), ethylenediaminetetraacetic acid (Sigma Aldrich), sodium phosphate (Sigma Aldrich), sodium sulfate (Sigma Aldrich), potassium bromide (Sigma Aldrich), ethanol amine (Sigma Aldrich), citric acid (Sigma Aldrich), sodium chloride (VWR).

U2OS cells were labeled with 10 μM EdU (Merck) for 30 min. Subsequently, cells were fixed with 1% formaldehyde (Merck) for 10 min, followed by quenching with 125 mM glycine (Merck) for 10 min. After two washing steps with PBS/1% BSA, cells were collected by scraping, followed by permeabilization in PBS/0.1% Triton X-100. Subsequently, cells were washed with PBS/1% BSA and subjected to the Click-iT reaction in a solution containing 10 mM sodium ascorbate (Merck), 0.1 mM azide-PEG₃-biotin conjugate (Merck) and 2 mM copper sulfate (Merck) for 30 min at room temperature. Cells were then washed twice in PBS/1% BSA, lysed in 10 mM Tris-HCl pH 8.0, 140 mM NaCl, 1% Triton X-100, 0.1% sodium deoxycholate, 0.1% SDS, supplemented with SIGMAFAST protease inhibitor cocktail, and sonicated using a Bioruptor (Diagenode). Lysates were cleared by centrifugation for 45 min at 4 °C in a table-top centrifuge and subjected to streptavidin-agarose beads (Thermo Fisher Scientific) overnight at 4 °C. The next day, beads were washed five times in PBS/1% BSA and de-crosslinking was carried out for 30 min in NuPAGE® LDS Sample Buffer at 95 °C. For protein detection, samples were subjected to SDS-PAGE and immunoblotted with relevant antibodies (Supplementary Table 2).