Synthesis and application of a new cleavable linker for "click"-based affinity chromatography.
A new chemically-cleavable linker has been synthesised for the affinity-independent elution of biomolecules by classical affinity chromatography. This azo-based linker is shown to couple efficiently with "click" derivatised ligands such as biotin propargyl amide through a copper(I)-catalysed Huisgen 1,3-dipolar cycloaddition reaction. Binding to Affi-Gel matrices displaying ligands coupled to the new linker is both efficient and selective. The captured material may be readily released from the resin upon treatment with sodium dithionite. These mild elution conditions have allowed for the efficient isolation of the affinity partner from complex protein mixtures such as those found in fetal bovine serum.
Landi
Felicetta
F
School of Chemistry, The University of Edinburgh, Kings Buildings, West Mains Road, Edinburgh, UKEH9 3JJ.
Johansson
Conny M
CM
Campopiano
Dominic J
DJ
Hulme
Alison N
AN
D000818:Animals; D001391:Azo Compounds / Q000138:chemical synthesis* / Q000737:chemistry; D001710:Biotin / Q000737:chemistry; D002417:Cattle; D002846:Chromatography, Affinity / Q000379:methods*; D008024:Ligands; D015394:Molecular Structure; D011506:Proteins / Q000302:isolation & purification*; D044967:Serum / Q000737:chemistry
D001391:Azo Compounds; D008024:Ligands; D011506:Proteins; D001710:Biotin