Gene Cloning and Characterization ofL-Ribulose 3-epimerase fromMesorhizobium lotiand Its Application to Rare Sugar Production

Abstract

A gene encoding L-ribulose 3-epimerase (L-RE) from Mesorhizobium loti, an important enzyme for rare sugar production by the Izumoring strategy, was cloned and overexpressed. The enzyme showed highest activity toward L-ribulose (230 U/mg) among keto-pentoses and keto-hexoses. This is the first report on a ketose 3-epimerase showing highest activity toward keto-pentose. The optimum enzyme reaction conditions for L-RE were determined to be sodium phosphate buffer (pH 8.0) at 60 °C. The enzyme showed of higher maximum reaction a rate (416 U/mg) and catalytic efficiency (43 M(-1) min(-1)) for L-ribulose than other known ketose 3-epimerases. It was able to produce L-xylulose efficiently from ribitol in two-step reactions. In the end, 7.2 g of L-xylulose was obtained from 20 g of ribitol via L-ribulose at a yield of 36%.

Bioconversion of d-glucose to d-psicose with immobilized d-xylose isomerase and d-psicose 3-epimerase on Saccharomyces cerevisiae spores

“…ST-24, Agrobacterium tumefaciens, Rhodobacter sphaeroides SK011, Clostridium cellulolyticum H10, Clostridium scindens 35704, Clostridium bolteae, Treponema primitia ZAS-1, Ruminococcus sp., Mesorhizobium loti, Desmospora sp, and Clostridium sp. have been characterized and employed for d-psicose synthesis [9,11,12,20,21,32,33,[36][37][38][39][40]. DTEases from A. tumefaciens, C. cellulolyticum H10, C. scindens 35704, C. bolteae, T. primitia, Ruminococcus sp., Desmospora sp, and Clostridium sp.…”

Advances in the enzymatic production of l-hexoses

“…The epimerase isolated from P. cichorii not only displayed 20 % of the relative activity toward L-psicose, compared to the optimum substrate D-tagatose, but also showed trace activity toward L-tagatose, L-sorbose, and Lfructose (Itoh et al 1994). The epimerase isolated from M. loti exhibited specific activities of 31, 10, and 5.5 U mg −1 toward L-psicose, L-fructose, and L-tagatose, respectively, and by comparison, showed a specific activity of 230 U mg −1 toward the optimum substrate L-ribulose and only trace activity toward L-tagatose (Uechi et al 2013b).…”

“…CAG317 , and Treponema primitia (Zhang et al 2016). Finally, a ketose 3-epimerase with the optimum substrate of L-ribulose, named L-ribulose 3-epimerase (EC 5.1.3.31), was first identified from Mesorhizobium loti (Uechi et al 2013b).…”

“…These enzymes are also used for the production of deoxyketohexoses (Gullapalli et al 2010;Rao et al 2009), methyl-ketohexoses (Jones et al 2008;Rao et al 2008), and some other rare sugars, including D-sorbose (Itoh et al 1995), L-xylulose (Uechi et al 2013b), L-tagatose, and L-fructose (Itoh and Izumori 1996). Although they exhibit broad substrate specificity toward various ketoses, the activities of many ketose 3-epimerases toward L-sugars have not been measured, probably due to the difficulty in obtaining Lsugars as substrates commercially.…”

“…Although they exhibit broad substrate specificity toward various ketoses, the activities of many ketose 3-epimerases toward L-sugars have not been measured, probably due to the difficulty in obtaining Lsugars as substrates commercially. Only the D-psicose 3-epimerase from P. cichorii (Izumori et al 1993) and the Lribulose 3-epimerase from M. loti (Uechi et al 2013b), identified by the Rare Sugar Research Center at Kagawa University, have been used to measure the epimerization activity toward L-ketohexoses. The epimerase isolated from P. cichorii not only displayed 20 % of the relative activity toward L-psicose, compared to the optimum substrate D-tagatose, but also showed trace activity toward L-tagatose, L-sorbose, and Lfructose (Itoh et al 1994).…”