期刊
CHEMBIOCHEM
卷 14, 期 18, 页码 2447-2455出版社
WILEY-V C H VERLAG GMBH
DOI: 10.1002/cbic.201300551
关键词
ABPP; analytical methods; click chemistry; CuAAC; proteomics
资金
- Deutsche Forschungsgemeinschaft (CIPSM) [SFB749, SFB1035]
- European Research Council (ERC) [259024]
- TUM Graduate School (Faculty Center Chemistry)
- European Research Council (ERC) [259024] Funding Source: European Research Council (ERC)
Copper-catalysed alkyne-azide 1,3-dipolar cycloaddition (CuAAC) is the predominantly used bioconjugation method in the field of activity-based protein profiling (ABPP). Several limitations, however, including conversion efficiency, protein denaturation and buffer compatibility, restrict the scope of established procedures. We introduce an ABPP customised click methodology based on refined CuAAC conditions together with new accelerating copper ligands. A screen of several triazole compounds revealed the cationic quaternary {3-[4-({bis[(1-tert-butyl-1H-1,2,3-triazol-4-yl)methyl]amino}methyl)-1H-1,2,3-triazol-1-yl]propyl}trimethylammonium trifluoroacetate (TABTA) to be a superior ligand. TABTA exhibited excellent in vitro conjugation kinetics and optimal ABPP labelling activity while almost exclusively preserving the native protein fold. The application of this CuAAC-promoting system is amenable to existing protocols with minimal perturbations and is even compatible with previously unusable buffer systems such as TrisHCl.
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