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5, purified uS10-3HA-6His were denatured with 1% SDS, reduced with 10 mM dithiothreitol (DTT) at 70 C for 10 min, and alkylated with 15 mM iodoacetamide (IAA) for another 15 min at room temperature in the dark, followed by on-beads tryptic digestion based on a single-pot solid-phase-enhanced sample preparation (SP3) method (Mller et al, 2020) using a KingFisher APEX equipped with a 96 Combi head (Thermo Fisher Scientific)

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