Reference : A spectrophotometric assay of D-glucuronate based on Escherichia coli uronate isomera...
Scientific journals : Article
Life sciences : Biochemistry, biophysics & molecular biology
http://hdl.handle.net/10993/13764
A spectrophotometric assay of D-glucuronate based on Escherichia coli uronate isomerase and mannonate dehydrogenase
English
Linster, Carole mailto [University of Luxembourg > Luxembourg Centre for Systems Biomedicine (LCSB) > >]
Van Schaftingen, Emile [> >]
Oct-2004
Protein Expression & Purification
Academic Press
37
2
352-60
Yes (verified by ORBilu)
International
1046-5928
1096-0279
Orlando
FL
[en] Escherichia coli uronate isomerase and mannonate dehydrogenase were overexpressed in E. coli BL21(DE3)pLysS cells and purified to near-homogeneity. The kinetic properties of the two enzymes were investigated. The isomerase was found to be inhibited by EDTA and to be stimulated by Zn(2+), Co(2+), and Mn(2+), but not by Mg(2+) or Ca(2+). Both enzymes were used to develop a sensitive spectrophotometric assay, in which D-glucuronate is converted to D-mannonate with concomitant oxidation of NADH to NAD(+). The sensitivity of this assay permits the detection of less than 1 nmol D-glucuronate. This assay can also be used to determine the concentration of beta-glucuronides and glucuronate 1-phosphate after enzymatic hydrolysis of these compounds with beta-glucuronidase or alkaline phosphatase.
http://hdl.handle.net/10993/13764
10.1016/j.pep.2004.06.015

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