Publications

• X-ray structure of Arthrobacter globiformis M30 ketose 3-epimerase for the production of D-allulose from D-fructose.

  Yoshida H., Yoshihara A., Gullapalli P.K., Ohtani K., Akimitsu K., Izumori K., Kamitori S.

  The X-ray structure of ketose 3-epimerase from Arthrobacter globiformis M30, which was previously reported to be a D-allulose 3-epimerase (AgD-AE), was determined at 1.96 Å resolution. The crystal belonged to the hexagonal space group P6₅22, with unit-cell parameters a = b = 103.98, c = ... >> More

  The X-ray structure of ketose 3-epimerase from Arthrobacter globiformis M30, which was previously reported to be a D-allulose 3-epimerase (AgD-AE), was determined at 1.96 Å resolution. The crystal belonged to the hexagonal space group P6₅22, with unit-cell parameters a = b = 103.98, c = 256.53 Å. The structure was solved by molecular replacement using the structure of Mesorhizobium loti L-ribulose 3-epimerase (MlL-RE), which has 41% sequence identity, as a search model. A hexagonal crystal contained two molecules in the asymmetric unit, and AgD-AE formed a homotetramer with twofold symmetry. The overall structure of AgD-AE was more similar to that of MlL-RE than to the known structures of D-psicose (alternative name D-allulose) 3-epimerases (D-PEs or D-AEs), although AgD-AE and MlL-RE have different substrate specificities. Both AgD-AE and MlL-RE have long helices in the C-terminal region that would contribute to the stability of the homotetramer. AgD-AE showed higher enzymatic activity for L-ribulose than D-allulose; however, AgD-AE is stable and is a unique useful enzyme for the production of D-allulose from D-fructose. << Less

  Acta Crystallogr. F Struct. Biol. Commun. 74:669-676(2018) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• TM0416 is a hyperthermophilic promiscuous non-phosphorylated sugar isomerase that catalyzes various C5 and C6 epimerization reactions.

  Shin S.M., Cao T.P., Choi J.M., Kim S.B., Lee S.J., Lee S.H., Lee D.W.

  There is currently little information on nonphosphorylated sugar epimerases, which are of potential interest for producing rare sugars. We found a gene (the TM0416 gene) encoding a putative d-tagatose-3-epimerase-related protein from the hyperthermophilic bacterium <i>Thermotoga maritima</i> We ov ... >> More

  There is currently little information on nonphosphorylated sugar epimerases, which are of potential interest for producing rare sugars. We found a gene (the TM0416 gene) encoding a putative d-tagatose-3-epimerase-related protein from the hyperthermophilic bacterium <i>Thermotoga maritima</i> We overexpressed the TM0416 gene in <i>Escherichia coli</i> and purified the resulting recombinant protein for detailed characterization. Amino acid sequence alignment and a structural similarity search revealed that TM0416 is a putative nonphosphorylated sugar epimerase. The recombinant enzyme exhibited maximal C-3 epimerization of l-ribulose to l-xylulose at ∼80°C and pH 7 in the presence of 1 mM Mn²⁺ In addition, this enzyme showed unusually high activity for the epimerization of d-tagatose to d-sorbose, with a conversion yield of 20% after 6 h at 80°C. Remarkably, the enzyme catalyzed the isomerization of d-erythrose or d-threose to d-erythrulose significantly, with conversion yields of 71% and 54.5%, respectively, after 6 h at 80°C at pH 7. To further investigate the substrate specificity of TM0416, we determined its crystal structures in complex with divalent metal ions and l-erythrulose at resolutions of 1.5 and 1.6 Å. Detailed inspection of the structural features and biochemical data clearly demonstrated that this metalloenzyme, with a freely accessible substrate-binding site and neighboring hydrophobic residues, exhibits different and promiscuous substrate preferences, compared with its mesophilic counterparts. Therefore, this study suggests that TM0416 can be functionally classified as a novel type of l-ribulose 3-epimerase (R3E) with d-erythrose isomerase activity.<b>IMPORTANCE</b> Rare sugars, which occur naturally in small amounts, have attracted considerable attention in the food and drug industries. However, there is little information on nonphosphorylated sugar epimerases, which might potentially be applied for the production of rare sugars. This study describes the characterization and functional annotation of a putative nonphosphorylated sugar 3-epimerase from a hyperthermophilic bacterium. Furthermore, we determined its crystal structures in complex with divalent metal ions and l-erythrulose, highlighting its metal-dependent, bifunctional, sugar-isomerizing activity. This hyperthermophilic R3E exhibited d-erythrose/d-threose isomerase activity, with structural features near the substrate-binding site distinct from those of its mesophilic counterparts. Moreover, this metalloenzyme showed unusually high activity for the epimerization of d-tagatose to d-sorbose at 70°C. Therefore, TM0416 can be functionally classified as a novel type of promiscuous R3E with a potential for the production of rare sugars for the food and pharmaceutical industries. << Less

  Appl. Environ. Microbiol. 0:0-0(2017) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• Gene cloning and characterization of L-ribulose 3-epimerase from Mesorhizobium loti and its application to rare sugar production.

  Uechi K., Takata G., Fukai Y., Yoshihara A., Morimoto K.

  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 repo ... >> More

  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%. << Less

  Biosci. Biotechnol. Biochem. 77:511-515(2013) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.