1xym
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THE ROLE OF THE DIVALENT METAL ION IN SUGAR BINDING, RING OPENING, AND ISOMERIZATION BY D-XYLOSE ISOMERASE: REPLACEMENT OF A CATALYTIC METAL BY AN AMINO-ACID
OverviewOverview
The distinct roles of the two magnesium ions essential to the activity of, D-xylose isomerase from Streptomyces olivochromogenes were examined. The, enzyme-magnesium complex was isolated, and the stoichiometry of cation, binding determined by neutron activation analysis to be 2 mol of magnesium, per mole of enzyme. A plot of Mg2+ added versus Mg2+ bound to enzyme is, consistent with apparent KD values of < or = 0.5-1.0 mM for one Mg2+ and <, or = 2-5 mM for the second. A site-directed mutant of D-xylose isomerase, was designed to remove the tighter, tetracoordinated magnesium binding, site (site 1, Mg-1); Glu180 was replaced with Lys180. The stoichiometry of, metal binding to this mutant, E180K, is 1 mol of magnesium per mole of, enzyme. Ring-opening assays with 1-thioglucose (H2S released upon ring, opening) show E180K catalyzes the opening of the sugar ring at 20% the, rate of the wild-type, but E180K does not catalyze isomerization of, glucose to fructose. Thus, the magnesium bound to Glu180 is essential for, isomerization but not essential for ring opening. The X-ray, crystallographic structures of E180K in the absence of magnesium and in, the presence and absence of 250 mM glucose were obtained to 1.8-A, resolution and refined to R factors of 17.7% and 19.7%, respectively. The, wild-type and both E180K structures show no significant structural, differences, except the epsilon-amino group of Lys180, which occupies the, position usually occupied by the Mg-1.(ABSTRACT TRUNCATED AT 250 WORDS)
About this StructureAbout this Structure
1XYM is a Single protein structure of sequence from Streptomyces olivochromogenes with GLO, MG and OH as ligands. Active as Xylose isomerase, with EC number 5.3.1.5 Full crystallographic information is available from OCA.
ReferenceReference
Role of the divalent metal ion in sugar binding, ring opening, and isomerization by D-xylose isomerase: replacement of a catalytic metal by an amino acid., Allen KN, Lavie A, Glasfeld A, Tanada TN, Gerrity DP, Carlson SC, Farber GK, Petsko GA, Ringe D, Biochemistry. 1994 Feb 15;33(6):1488-94. PMID:7906142
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