Xylose Isomerase in Substrate and Inhibitor Michaelis States:  Atomic Resolution Studies of a Metal-Mediated Hydride Shift^,

Abstract

Xylose isomerase (E.C. 5.3.1.5) catalyzes the interconversion of aldose and ketose sugars and has an absolute requirement for two divalent cations at its active site to drive the hydride transfer rates of sugar isomerization. Evidence suggests some degree of metal movement at the second metal site, although how this movement may affect catalysis is unknown. The 0.95 A resolution structure of the xylitol-inhibited enzyme presented here suggests three alternative positions for the second metal ion, only one of which appears positioned in a catalytically competent manner. To complete the reaction, an active site hydroxyl species appears appropriately positioned for hydrogen transfer, as evidenced by precise bonding distances. Conversely, the 0.98 A resolution structure of the enzyme with glucose bound in the alpha-pyranose state only shows one of the metal ion conformations at the second metal ion binding site, suggesting that the linear form of the sugar is required to promote the second and third metal ion conformations. The two structures suggest a strong degree of conformational flexibility at the active site, which seems required for catalysis and may explain the poor rate of turnover for this enzyme. Further, the pyranose structure implies that His53 may act as the initial acid responsible for ring opening of the sugar to the aldose form, an observation that has been difficult to establish in previous studies. The glucose ring also appears to display significant segmented disorder in a manner suggestive of ring opening, perhaps lending insight into means of enzyme destabilization of the ground state to promote catalysis. On the basis of these results, we propose a modified version of the bridged bimetallic mechanism for hydride transfer in the case of Streptomyces olivochromogenes xylose isomerase.

Enzymatic Characterization of 5-Methylthioribose 1-Phosphate Isomerase fromBacillus subtilis

“…These features of the reaction were similar to those of xylose isomerase, which adopts the hydride transfer mechanism. 18,24) These results suggest that B. subtilis MTR-1-P isomerase follows the hydride transfer mechanism in its isomerization reaction. An interesting point, unique to B. subtilis MTR-1-P isomerase, is that this enzyme catalyzed the reaction without the participation of a metal ion in the reaction (Table 1).…”

“…19,20) The other mechanism is the hydride transfer mechanism, in which a hydride on a carbon is directly transferred to the neighboring cationic carbon. 18,23,24) In the isomerase reaction of B. subtilis MTR-1-P isomerase, the product of the enzyme reaction in the presence of 99.9% D 2 O showed no increase in mass after the reaction, in contrast to the case with R5P isomerase (Table 2), and was virtually deuterium-free in NMR analysis (Fig. 4, inset).…”

Metalloenzyme-like catalyzed isomerizations of sugars by Lewis acid zeolites

“…The enzymatic isomerization step is believed to occur primarily at M2. First, a hydroxyl ion bound to M2 promotes deprotonation at O2 (17). Next, M2 forms a bidentate complex with both O1 and O2, withdrawing electron density from the C-O bonds and effectively polarizing the substrate.…”

Comparison of Homogeneous and Heterogeneous Catalysts for Glucose‐to‐Fructose Isomerization in Aqueous Media

“…[21] In all three cases, the active metal center (Lewis acid) coordinates to oxygen atoms in the first shell and has a covalently bonded À OH (Brønsted base) group. In addition, it has been shown that the glucose molecule binds to [Cr(H 2 O) 5 …”

“…Thus, the isomerization catalyst should be compatible with the subsequent dehydration reaction, which is performed by using a Brønsted-acid catalyst at relatively high temperatures (> 413 K). The aldose-to-ketose isomerization is used in large industrial applications, which use enzymatic or base-catalyzed reactions; [5] however, neither of these processes is compatible with the dehydration reaction when using a single reactor. Thus, the development of catalysts for this reaction in aqueous media is an important milestone in our ability to utilize biomass.…”