Summary
The separation of glycoproteins as glycoforms with specific degrees of glycosylation has been a problem until now. A technique involving boronate affinity chromatography has been developed to separate a heterogeneous sample of neoglycoprotein, chymotrypsin modified with maltose via reductive amination, into individual fractions with different degrees of glycosylation. Low-molecular-mass polyhydroxyl compounds, such as tris(hydroxymethyl)aminomethane (Tris), pentaerythritol, and triethanolamine proved efficient eluents due to their ability to form strong tridentate complexes with the boronate ligand. Compounds leading to either; too strong interaction with the boronate ligand (e. g. D-sorbitol and polyvinly alcohol), or too weak or no interaction (e. g. dextran) were not suitable eluents. The study provided the opportunity to probe further into the effect of glycosylation on the function of glycoproteins.
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References
Davis, B.G.J. Chem. Soc. Perkin Trans. 1999,1, 3215–3237.
Raju, T.S.; Lerner, L.; O'Connor, J.V.Biotechnol. Appl. Biochem. 1996,24, 191–194.
Stowell, C.P.; Lee, Y.C.Adv. Carbohydr. Chem. Biochem. 1980,37, 225–281.
Gonzales, R.; Thiem, J.Carbohydr. Res. 1999,317, 180–190.
Lee, Y.C.; Lee, R.T.Methods Enzymol. 1994,242, 3–123.
Rademacher, T.W.; Parekh, R.B.; Dwek, R.A.Ann. Rev. Biochem. 1988,57, 785–838.
Lee, R.T.; Lee, Y.C. InGlycosciences (Status and Perspectives): Gabius, H.; Gabius, S. Eds. Chapman & Hall, Weinheim,1997.
Weith, H.L.; Wiebers, J.L.; Gilham, P.T.Biochemistry,1970,9, 4396–4401.
Fulton, S.Boronate lignads in biochemical separations. Amicon Corporation, Danvers, MA, USA.,1981.
Bergold, A.; Scouten, W.H. InSolid phase biochemistry (analytical and synthetic methods): Scouten, W.H. Ed, John Wiley & Sons,1983.
Middle, F.A.; Bannister, A.; Bellingham, A.J.; Dean, P.D.G.Biochem. J. 1983,209, 771–779.
Williams, G.G.; Johnstone, A.P.; Bouriotis, V.; Dean, P.D.G.Biochem. Soc. Trans. 1981,9, 137–139.
Mazzeo, J.R.; Krull, I.S.Bio Chromatography,1989,4, 124–130.
Hagmen, J.H.; Kuehn, G.D. InMethods in Molecular Biology: Kenney, A.; Fowell, S. Eds, The Humana Press Inc. Totowa, NJ,vol. 11,1992.
Gerstner, J.A.; Londo, T.; Hunt, T.; Morris, J.; Pedroso, P.; Hamilton, P.R.Chem-Tech,1995,November, 27–32.
Bradford, M.M.Anal. Biochem. 1976,72, 248–254.
Scott, T.A.; Melvin, E.H.Anal. Chem. 1953,25, 1656–1661.
Schwartz, A.; Gary, G.R.Arch. Biochem. Biophys. 1977,181, 542–549.
Li, Y.C.; Linné Larsson, E.; Jungvid, H.; Galaev, I. Yu. Mattiasson, B.Bioseparation 2001,9, 315–323.
Li, Y.C.; Linné Larsson, E.; Jungvid, H.; Galaev, I.Yu.; Mattiasson, B.J. Chromatogr. A 2000,909, 137–145.
Zittle, C.Advan. Enzym. 1951,12, 493–496.
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Li, Y.C., Larsson, E.L., Jungvid, H. et al. Separation of neoglycoproteins with different degrees of glycosylation by boronate chromatography. Chromatographia 54, 213–217 (2001). https://doi.org/10.1007/BF02492248
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DOI: https://doi.org/10.1007/BF02492248