Preliminary Characterization of Xylose Reductase Partially Purified by Reversed Micelles from <i>Candida tropicalis</i> IEC5-ITV, an Indigenous Xylitol-Producing Strain

Abstract

Xylose reductase (EC 1.1.1.21) of Candida tropicalis IEC5-ITV, an indigenous xylitol-producing strain, was partially purified by reversed micelles and characterized, an 8.1 fold purification factor being obtained. The XR present in the crude extract exhibited its highest specific activity at pH 6.0 and 40˚C, while in that obtained by reverse micelles, this occurs at pH 6.0 and 30˚C. XR before and after extraction is stable within a range of 30˚C to 40˚C, pH 7 after one hour of incubation under these conditions. After two months' storage at -18˚C, the enzyme obtained by reverse micelles lost 76.60% specific activity. The estimated molecular weight by PAGE-SDS was 32.42 kD. K M for xylose was higher for the XR extracted by reverse micelles (0.026 M) than that obtained for the enzyme before extraction (0.0059 M), while K M for cofactor NADPH was lower after than before extraction (1.85 mM to 12.0 mM respectively). There was no activity with NADH as a cofactor. Variations in pH and temperature optima, as well as kinetic parameters before and after partial XR purification by reverse micelles are probably due to an alteration in enzyme molecule structure caused by the solvents used during extraction.

Effects of agitation rate and dissolved oxygen on xylose reductase activity during xylitol production at bioreactor scale

“…Subsequently, 100 µL of a 0.5 M xylose solution were added and the absorbance was measured every 6 s for 60 s. The XR unit (U) was defined as the amount of enzyme that catalyzes the reduction of 1 µmol of NADPH per min. The specific XR activity was expressed as a unit of the enzyme per mg of protein with Equation 1 (Cocotle-Ronzon et al, 2012).…”

“…Dissolved oxygen is one of the most important factors affecting xylitol production, especially when Candida is used for the bioconversion of D-xylose (Albuquerque et al, 2014). During the fermentation process, Candida metabolizes xylose into xylitol in the presence of key enzymes that participate in a metabolic pathway such as xylose reductase (XR; EC1.1.1.21); an intracellular enzyme located in the cytoplasm, which has the ability to reduce xylose to xylitol in the first phase of the metabolic pathway in the presence of NADPH as a cofactor (Cocotle-Ronzon et al, 2012). The XR activity has also special scientific attention for its application in the fermentation of lignocellulosic substrates for the production of ethanol and xylitol (Zhang et al, 2019).…”

Modeling of the xylitol production from sugarcane bagasse by immobilized cells

“…9,10 Cell immobilization is a strategy to improve bioconversion since it allows microorganisms to be not affected by the presence of toxic compounds in the environment and can also be reused in various experimental tests. [11][12][13] Different materials have been used for immobilization such as calcium alginate, [14][15][16] sugarcane bagasse, 17 porous glass 18 and hydrogel of polyvinyl alcohol. 19 Among them, calcium alginate is one of the most accepted in the food industry, since it has a low cost and it does not affect the capability or activity of microorganisms.…”

Xylitol production by Saccharomyces cerevisiae overexpressing different xylose reductases using non-detoxified hemicellulosic hydrolysate of corncob

“…requirements of high pressure, temperature, expensive catalyst, and extensive downstream operations (Leathers 2003 ). Microorganisms convert xylose to xylitol by the cofactor-dependent xylose reductase (XR or Xyl1 ) enzyme (Saha 2003 ; Cocotle-Ronzon et al 2012 ). The majority of essential cofactor NADPH/NADH for the XR activity is regenerated through pentose phosphate pathway (PPP) (Bengtsson et al 2009 ; Schwartz et al 2012 ).…”