Traceless Cleavage of Protein–Biotin Conjugates under Biologically Compatible Conditions |
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| Authors: | Dr Joseph Cowell Dr Matthew Buck Dr Ali H Essa Rebecca Clarke Prof Waldemar Vollmer Dr Daniela Vollmer Dr Catharien M Hilkens Prof John D Isaacs Dr Michael J Hall Dr Joe Gray |
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| Affiliation: | 1. School of Chemistry, Newcastle University, Newcastle upon Tyne, UK;2. Musculoskeletal Research Group, Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK;3. Department of Chemistry, College of Science, University of Basrah, Basrah, Iraq;4. Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne, UK |
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| Abstract: | Biotinylation of amines is widely used to conjugate biomolecules, but either the resulting label is non‐removable or its removal leaves a tag on the molecule of interest, thus affecting downstream processes. We present here a set of reagents (RevAmines) that allow traceless, reversible biotinylation under biologically compatible, mild conditions. Release following avidin‐based capture is achieved through the cleavage of a (2‐(alkylsulfonyl)ethyl) carbamate linker under mild conditions (200 mm ammonium bicarbonate, pH 8, 16–24 h, room temperature) that regenerates the unmodified amine. The capture and release of biotinylated proteins and peptides from neutravidin, fluorescent labelling through reversible biotinylation at the cell surface and the selective enrichment of proteins from bacterial periplasm are demonstrated. The tags are easily prepared, stable and offer the potential for future application in proteomics, activity‐based protein profiling, affinity chromatography and bio‐molecule tagging and purification. |
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| Keywords: | affinity purification protein modifications proteomics reversible biotinylation traceless cleavage |
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