Abstract
Affinity chromatography, the method of purifying target proteins from complex mixtures using immobilized affinity ligands on chromatographic supports, is perhaps the most common of all affinity techniques (1–3). Many affinity chromatography systems are comprised of activated supports requiring direct ligand-coupling procedures that can be complex, time-consuming, and result in low- or variable-capacity columns supporting immobilized ligands with poor activity (2–5). We describe here a method that improves this technology by using a small-molecule based chemical affinity technology (6) to quickly and easily prepare high-capacity affinity columns supporting functionally active capture ligands for purifying proteins from crude mixtures (7,8). This innovation is based on the specific interaction between two, non-biological, small molecules, phenyl-diboronic acid (PDBA) and salicylhydroxamic acid (SHA) (see Fig. 1).
PDBA:SHA Chemical Affinity System. The reaction of phenyldiboronic acid (PDBA) with salicylhydroxamic acid (SHA). The SHA is covalently anchored to the surface of crosslinked agarose chromatography media and a PDBA derivative is conjugated to a protein of choice.
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Hughes, K.A., Wiley, J.P. (2003). Small-Molecule Affinity-Based Matrices for Rapid Protein Purification. In: Vaillancourt, P.E. (eds) E. coliGene Expression Protocols. Methods in Molecular Biology™, vol 205. Humana Press. https://doi.org/10.1385/1-59259-301-1:215
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DOI: https://doi.org/10.1385/1-59259-301-1:215
Publisher Name: Humana Press
Print ISBN: 978-1-58829-008-3
Online ISBN: 978-1-59259-301-9
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