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Chromatography

Chromatography

In principle, the specific interaction of cyclodextrins with aromatic amino-acid side-chains of proteins can be exploited in chromatographic applications. Proteins modified with synthetic fluorophores1 or prenyl groups2 show strong affinity towards β-CD-modified Sepharose, allowing the chromatographic separation of these modified proteins from unmodified counterparts.

We are currently developing and investigating various cyclodextrin-based chromatographic materials, using silica3 or polysaccharides beads as solid supports for HPLC or FPLC systems. These materials are tested for their performance in separating various proteins in native or partially denatured conformations, depending on the relative exposure of aromatic acid side chains to the aqueous solvent. This form of chromatography could represent a better alternative to classical hydrophobic interaction (HIC), which typically requires harsher conditions (high salt concentration for binding or organic solvents for elution).

References

  • Nguyen, T., Joshi, N.S., Francis, M.B. 2006. An affinity-based method for the purification of fluorescently-labeled biomolecules. Bioconjugate Chem. 17, 869−872.
  • Chung, JA, Wollack, JW, Hovlid, ML, Okesli, A, Chen, Y, Mueller, JD, Distefano, MD, Taton, TA, 2009. Purification of prenylated proteins by affinity chromatography on cyclodextrin-modified agarose. Anal Biochem. 386, 1-8.
  • Nielsen, R. B. H., Nielsen, J. L., & Larsen, K. L. (2010). Distribution and accessibility of cyclodextrins covalently bound onto silica gel. J Incl Phenom Macroc Chem, 67, 399-405.