PMID: 2505925Jun 15, 1989Paper

Purification of endo-N-acetyl-beta-D-glucosaminidase H by substrate-affinity chromatography

Carbohydrate Research
U F GreberK Mosbach

Abstract

Endo-N-acetyl-beta-D-glucosaminidase H (Endo H) was purified to homogeneity (3000-fold) from a culture filtrate to Streptomyces plicatus. The key step was substrate-affinity chromatography, which afforded a 1000-fold purification and yielded a protease- and exoglycosidase-free preparation of Endo H. Proteins from the crude sample were applied to the substrate-affinity column, consisting of yeast-invertase glycopeptides bound to Sepharose-immobilized concanavalin A. After washing off the unbound proteins, Endo H was quantitatively eluted by methyl alpha-D-mannopyranoside. Various conditions were tested to achieve an optimal binding of Endo H to this substrate-affinity gel. After substrate-affinity chromatography, Endo H was separated from the coeluted gylcopeptide substrate and some protein impurities by gel filtration and hydrophobic chromatography.

References

Jul 1, 1989·European Journal of Clinical Microbiology & Infectious Diseases : Official Publication of the European Society of Clinical Microbiology·H C Neu, N X Chin
Jan 1, 1985·Annual Review of Biochemistry·R Kornfeld, S Kornfeld
Feb 1, 1967·Biochemistry·N P Neumann, J O Lampen
Apr 1, 1966·Analytical Biochemistry·R Gatt, E R Berman
Nov 1, 1984·Archives of Biochemistry and Biophysics·S BarbarićP Mildner
Sep 1, 1981·Analytical Biochemistry·M NummiV Raunio

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