Xylulose dehydrogenase

The reactivity of xylitol in the polyol dehydrogenase reactions has been extensively studied for its relations to pentosuria. Xylitol was found to be oxidized in two ways (a) to L-xylulose by a highly specific NADP-requiring dehydrogenase and (b) to D-xylulose by a NAD-linked enzyme with lesser substrate specificity ... 

Hll. Hollman, S., and Touster, O., The L-xylulose-xylitol enzyme and other polyol dehydrogenases of guinea pig liver mitochondria. ]. Biol. Chem. 225, 87-102 (1957). 

D-3) Essential pentosuria (deficiency of xylitol dehydrogenase). A benign accumulation of xylulose develops, which may be confused with glucose when detected in the urine. 

L-Xylulose reductase (xylitol dehydrogenase) is deficient in essential pentosuria. L-Xylulose (a pentose) appears in the urine and gives a positive reducing-sugar test. The condition is benign. 

Fig. 5. Assimilation of D-xylose, L-arabinose and D-arabinose. In yeasts and fungi, pentoses are assimilated by way of oxidoreductases. D-xylose. L-arabinose and D-arabinose are each reduced to their respective polyols by aldose reductases, designated here as Xor, Lar and Dar. Both D-xylose and L-xylose are reduced to xylitol, which is symmetrical. D-xylose and L-arabinose are the forms normally found in nature. D- and L-arabitol dehydrogenases (Dad and Lad) form D- and L-xylulose, respectively. D- and L-Xylitol dehydrogenase (Dxd and Lxd) mediate the formation of D- and L-xylulose from xylitol...

Yeasts and bacteria metabolize xylose by following sHghtly different pathways as showing in Fig. 1. Yeasts rely on xylose reductase and xyHtol dehydrogenase, but bacteria rely on xylose isomerase, to convert xylose to xylulose [2, 3]. Although the Saccharomyces yeasts as well as other fermentative yeasts are not able to ferment xylose, Saccharomyces yeasts are able to ferment xylulose to ethanol [4]. Furthermore, they are also able to ferment xylose when a bacterial xylose isomerase is present in the medium [5]. This indicates that Saccharomyces yeasts lack only the enzymes for the conversion of xylose to xylulose. 

The Pichia xylitol dehydrogenase gene chosen by us to be cloned into the Saccharomyces yeast is known to produce a xylitol dehydrogenase that catalyzes a reversible reaction between xylitol and xylulose as shown in Fig. 1 but favors the formation of xylitol rather than xylulose. Thus, an extremely strong xylulokinase activity will help to direct the carbon flux towards the production of ethanol rather than the formation of byproduct xylitol. 

Ethanol fermentation from xylose by yeasts can be divided into four distinctive steps. The first step is the reduction of xylose to xylitol mediated by NADPH/ NADH-linked xylose reductase (XR). This is followed by the oxidation of xylitol to xylulose, mediated by NAD-linked xylitol dehydrogenase (XDH). Xylulose-5-phosphate, the key intermediate, is generated from the phosphorylation of xylulose by xylulose kinase. Xylulose-5-phosphate is then channeled into the pentose phosphate pathway for further metabolism (Fig. 9). 

Xylitol dehydrogenase converts xylitol to the 2-ketopentose xylulose and the tetrameric enzyme from Galactocandida mastotermitis has been shown to possess one essential Zn " per monomer. As expected, binding is ordered with the cofactor binding first however, binding of carbohydrate is so weak that a Theorell-Chance kinetic mechanism obtains (he. one in which there is a bimolecular reaction between E.NAD and xylitol, without detectable E.NAD ". xylitol or E.NADEI.xylulose complexes). 

In fungi, xylose is reduced to xylitol by NADH- or NADPH-dependent xylose reductase (XR) and thereafter is oxidized to xylulose by NAD -dependent xylitol dehydrogenase (XDH). The xylulose is phosphorylated, channeled into the pentose phosphate pathway [3]. XR of most fungi, including most yeasts, prefers NADPH to NADH. Because of the cofactor preference of XR (NADPH) and XDH (NAD, redox imbalance occurs under anaerobic condition [4]. Therefore, the oxygen-limited rather than anaerobic condition is ideal for bioconversion of xylose to ethanol, so that the accumulated reduced cofactor can be oxidized to reach redox balance. A critical level of oxygen should exist for the highest ethanol yield and productivity. 

C. guilliermondii produces the enzyme D-xylose reductase which catalyses a reaction where the proton carrier NADPH donates a hydrogen atom to D-xylose, and D-xylose is converted to xylitol as seen in Fig. 1. The xylitol can then be converted to D-xylulose, catalyzed by xylitol dehydrogenase, which is utilized in central metabolism [3]. Under semi-aerobic conditions, xylitol accumulation is favored compared to anaerobic or aerobic conditions. Under anaerobic conditions, the ratio of NAD(P)H to NAD(P) is low, and NAD(P)H is required for xylitol production. Under aerobic conditions, excess oxygen allows oxidation of NADH to NAD", and a resulting high NAD /NADH ratio results in a faster xylitol conversion rate to D-xylulose, eliminating the accumulation of xylitol [4, 5]. 

The xylose reductase (XR) catalyzes the first step of a fungal pathway that allows certain organisms to metabolize xylose, such as Candida boidinii [6], Candida guilliermondii [7], Candida tmpicalis [8], Candida parapsilosis [9], and Debaryomyces hansenii [10]. After the reduction of xylose to xylitol by XR in a manner that can utilize nicotinamide adenine dinucleotide (reduced form NADH) or nicotinamide adenine dinucleotide phosphate (reduced form NADPH), xylitol is re-oxidized to xylulose by xyUtol dehydrogenase, which is often specific for nicotinamide adenine dinucleotide (NAD) [11]. Xylulose is then phosphorylated. An efficient, pathway should recycle the cosubstrate such that there is no... 

In yeast and filamentous fimgi, xylose is converted to xylulose in two steps, where the first reaction is catalyzed by xylose reductase (XR) and the second by xylitol dehydrogenase, (XDH) (Fig. 1) [24]. Procaryotic organisms use a xylose isomerase (XI) to perform the conversion in one step [132]. 

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