Glucosamine Research Today is a free monthly online journal that collates and summarizes the latest research about Glucosamine, including details on chondroitin sulfate, uses, effects, benefits, arthritis.

Construction, purification, and functional characterization of His-tagged Candida albicans glucosamine-6-phosphate synthase expressed in Escherichia coli.

Olchowy J, Kur K, Sachadyn P, Milewski S

Department of Pharmaceutical Technology and Biochemistry, Gdańsk University of Technology, 11/12 Narutowicza St., 80-952 Gdańsk, Poland.

Expression plasmids containing recombinant genes encoding three His(6)-tagged versions of the enzyme, glucosamine-6-phosphate synthase from Candida albicans, were constructed and overexpressed in Escherichia coli. The gene products were purified by metal-affinity chromatography to near homogeneity with 77-80% yield and characterized in terms of size and enzymatic properties. Presence of oligohistidyl tags at either of two ends did not affect enzyme quarternary structure but strongly influenced its catalytic activity. The His6-N-tagged enzyme completely lost an ability of glucosamine-6-phosphate formation and amidohydrolase activity but retained the hexosephosphate-isomerising activity. On the other hand, two His6-C-tagged versions of glucosamine-6-phosphate synthase exhibited amidohydrolase activity almost equal to that of the wild-type enzyme but only 18% of its hexosephosphate-isomerising activity and about 1.5% of the synthetic activity.

Published 3 April 2006 in Protein Expr Purif, 46(2): 309-15.
Full-text of this article is available online (may require subscription).

© 2004-2008 Glucosamine Research Today. All Rights Reserved.