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X-Ray Crystallographic Analysis of D-Xylose Isomerase-Catalyzed Isomerization of (R)-Glyceraldehyde
[XYLA_STRRU] Involved in D-xylose catabolism.
Publication Abstract from PubMed
D-Xylose isomerase (XI) and triosephosphate isomerase (TIM) catalyze the aldose-ketose isomerization reactions of D-xylose and D-glyceraldehyde 3-phosphate (DGAP), respectively. D-Glyceraldehyde (DGA) is the triose fragment common to the substrates for XI and TIM. The XI-catalyzed isomerization of DGA to give dihydroxyacetone (DHA) in D<sub>2</sub>O was monitored by <sup>1</sup>H NMR spectroscopy and <i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub> = 0.034 M<sup>-1</sup> s<sup>-1</sup> was determined for this isomerization at pD 7.0. This is similar to <i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub> = 0.017 M<sup>-1</sup> s<sup>-1</sup> for the TIM-catalyzed carbon deprotonation reaction of DGA in D<sub>2</sub>O at pD 7.0 [Amyes, T. L.; O'Donoghue, A. C. and Richard J. P. (2001) <i>J. Am. Chem. Soc. 123</i>, 11325-11326]. The much larger activation barrier for XI-catalyzed isomerization of D-xylose (<i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub> = 490 M<sup>-1</sup> s<sup>-1</sup>) than for the TIM-catalyzed isomerization of DGAP (<i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub> = 9.6 x 10<sup>6</sup> M<sup>-1</sup> s<sup>-1</sup>) is due to: (i) The larger barrier to conversion of cyclic D-xylose to the reactive linear sugar (5.4 kcal/mol) than for conversion of DGAP hydrate to the free aldehyde (1.7 kcal/mol). (ii) The smaller intrinsic binding energy [Jencks, W. P. (1975) <i>Adv. Enzymol. Relat. Areas Mol. Biol. 43</i>, 219-410] of the terminal ethylene glycol fragment of D-xylose (9.3 kcal/mol) than of the phosphodianion group of DGAP (ca. 12 kcal/mol). The XI catalyzed isomerization of DGA in D<sub>2</sub>O at pD 7.0 gives a 90% yield of [1-<sup>1</sup>H]-DHA and a 10% yield of [1-<sup>2</sup>H]-DHA, the product of isomerization with deuterium incorporation from solvent D<sub>2</sub>O. By comparison, the transfer of <sup>3</sup>H from labeled hexose substrate to solvent is observed only once in every 10<sup>9</sup> turnovers for the XI-catalyzed isomerization of [2-<sup>3</sup>H]-glucose in H<sub>2</sub>O [Allen, K. N., Lavie, A., Farber, G. K., Glasfeld, A., Petsko, G. A., and Ringe, D. (1994), <i>Biochemistry 33</i>, 1481-1487]. We propose that truncation of the terminal ethylene glycol fragment of D-xylose to give DGA results in a large decrease in the rate of XI-catalyzed isomerization with hydride transfer compared with that for proton transfer. An ultra-high resolution (0.97 A) X-ray crystal structure was determined for the complex obtained by soaking crystals of XI with 50 mM DGA. The triose binds to XI as the unreactive hydrate, but ligand binding induces metal cofactor movement and conformational changes in active site residues similar to those observed for XI-sugar complexes.
Binding Energy and Catalysis by D-Xylose Isomerase: Kinetic, Product and X-Ray Crystallographic Analysis of Enzyme-Catalyzed Isomerization of (R)-Glyceraldehyde.,Toteva MM, Silvaggi NR, Allen KN, Richard JP Biochemistry. 2011 Oct 13. PMID:21995300
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.