Glycogen_NoPro Workflow (IMAGE)
Caption
The depicted workflow illustrates how non-proteinaceous substrates of ubiquitin are processed for detection via NoPro-clipping, a method which was developed in this study. Briefly, ubiquitin is clipped off its substrates via Ub-clipping using the bacterial enzyme BpJOS. This step converts ubiquitin modifications into diglycine remnants. Subsequently, a small peptide (=ClipTag) is attached to these diglycine modifications via the enzyme Sortase A. Together, these steps transform ubiquitinated non-proteins into peptide-modified species (=ClipTag-conjugates) that can be readily detected by mass-spectrometry-based proteomics workflows.
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WEHI
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