A novel synthetic chemistry approach to linkage-specific ubiquitin conjugation† †Electronic supplementary information (ESI) available: Full experimental details are provided. See DOI: 10.1039/c5ob00130g Click here for additional data file.

Mass Spectrometry LC-MS was performed on protein samples using a Thermo Scientific uPLC connected to MSQ Plus Single Quad Detector (SQD). Column: Hypersil Gold C4 1.9m 2.1 x 50 mm. Wavelength: 254 nm. Mobile Phase: 99:1 Water (0.1% formic acid): MeCN (0.1% formic acid) to 1:9 Water (0.1% formic acid): MeCN (0.1% formic acid) gradient over 4 min. Flow Rate: 0.3 mL/min. MS Mode: ES+. Scan Range: m/z = 500-2000. Scan time: 1.5 s. Data obtained in continuum mode. The electrospray source of the MS was operated with a capillary voltage of 3.5 kV and a cone voltage of 50 V. Nitrogen was used as the nebulizer and desolvation gas at a total flow of 600 L/h. Total mass spectra for protein samples were reconstructed from the ion series using the pre-installed ProMass software using default settings for large proteins in m/z range 500-1500.