1XYM

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

Summary for 1XYM

• Entry DOI: 10.2210/pdb1xym/pdb
• Descriptor: XYLOSE ISOMERASE, D-glucose, MAGNESIUM ION, ... (5 entities in total)
• Functional Keywords: isomerase(intramolecular oxidoreductase)
• Biological source: Streptomyces olivochromogenes
• Total number of polymer chains: 2
• Total formula weight: 86132.76

Authors

Allen, K.N.,Lavie, A.,Petsko, G.A.,Ringe, D. (deposition date: 1993-12-07, release date: 1994-05-31, Last modification date: 2024-02-14)

Primary citation

Allen, K.N.,Lavie, A.,Glasfeld, A.,Tanada, T.N.,Gerrity, D.P.,Carlson, S.C.,Farber, G.K.,Petsko, G.A.,Ringe, D.
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.
Biochemistry, 33:1488-1494, 1994

PubMed Abstract: 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)

PubMed: 7906142
DOI: 10.1021/bi00172a027

Experimental method

X-RAY DIFFRACTION (1.8 Å)