Mamoru Nishimoto - Academia.edu (original) (raw)
Papers by Mamoru Nishimoto
FEBS letters, Jan 23, 2016
In Ruminococcus albus, either 4-O-β-d-mannosyl-d-glucose phosphorylase (RaMP1) and β-(1,4)-mannoo... more In Ruminococcus albus, either 4-O-β-d-mannosyl-d-glucose phosphorylase (RaMP1) and β-(1,4)-mannooligosaccharide phosphorylase (RaMP2) belong to two subfamilies of glycoside hydrolase family 130. The two enzymes phosphorylate β-mannosidic linkages at the non-reducing ends of their substrates, and have substantially diverse substrate specificity. The differences in their mechanism of substrate binding are yet not fully elucidated. Here, we report the crystal structures of RaMP1 with/without 4-O-β-d-mannosyl-d-glucose and RaMP2 with/without β-(1,4)-mannobiose. The structures of the two enzymes differ at the +1 subsite of the substrate binding pocket. Three loops are proposed to determine the different substrate specificities. Loop 3 interacts with the bound oligosaccharide. In RaMP1, His245 of loop 3 forms a hydrogen-bond network with the substrate through a water molecule, and is indispensible for substrate binding. This article is protected by copyright. All rights reserved.
Journal of Applied Glycoscience, 2014
Journal of Applied Glycoscience, 2001
Journal of Applied Glycoscience, 2002
Journal of Applied Glycoscience, 2011
Xylanase [EC 3.2.1.8] hydrolyzes the main chain of xylan (the major component of hemicelluloses),... more Xylanase [EC 3.2.1.8] hydrolyzes the main chain of xylan (the major component of hemicelluloses), consisting of β-1,4 xylosyl linkages in an endowise manner, and was also studied as an industrial enzyme for the utilization of unused biomass and a substitute for chemical bleach in paper manufacturing. 1 5) The natural substrate, xylan, is typically used for the characterization of xylanase. However, it is diffi cult to compare the characters of the enzymes determined with its natural substrate, xylan, for the following reasons: 1) Xylan is a heterogeneous polymer often heterogeneously decorated with α-L-arabinofuranosyl and/or 4-methyl-α-D-glucuronopyranosyl residues depending on its origin and lot; 6 11) 2) Several xylanases showed severe substrate inhibition of xylan. 12) Previously, we performed kinetic experiments of several xylanases using p-nitrophenyl-βxylobioside (XX-pNP) as the substrate. In such cases, no substrate inhibition was observed, and their activity could be evaluated accurately and reproducibly. 13) Xylanases belonging to the glycoside hydrolase family 10 (GH10) 14 16) have often been reported to hydrolyze cellobiosyl substrates such as p-nitrophenyl-β-cellobioside (GG-pNP). The ratio of the catalytic effi ciencies on XX-pNP and GG-pNP
Journal of Applied Glycoscience, 2014
One-pot enzymatic production of nigerose was demonstrated from abundantly available sugar resourc... more One-pot enzymatic production of nigerose was demonstrated from abundantly available sugar resources, including maltose, cellobiose, sucrose and starch. (i) 319 mM nigerose was generated from 500 mM maltose by the combined actions of maltose phosphorylase and nigerose phosphorylase, which share the same β-D-glucose 1-phosphate, in the presence of phosphate. The yield was 62% based on the concentration of maltose as the starting material. (ii) 129 mM nigerose was produced from 250 mM cellobiose by cellobiose phosphorylase and nigerose phosphorylase in the presence of phosphate, in combination with the enzymatic pathway to convert α-D-glucose 1-phosphate to β-D-glucose 1-phosphate via D-glucose 6-phosphate by the combined actions of α-phosphoglucomutase and β-phosphoglucomutase, resulting in a yield of 52%. (iii) 350 mM nigerose was produced from 500 mM sucrose by substituting cellobiose phosphorylase with sucrose phosphorylase and adding xylose isomerase, giving a yield of 67%. (iv) 270 mM nigerose was generated from 100 mg/mL starch and 500 mM D-glucose by the concomitant actions of glycogen phosphorylase, isoamylase, α-phosphoglucomutase, β-phosphoglucomutase and nigerose phosphorylase, in the presence of phosphate. In addition, 280 mM 3-O-α-D-glucopyranosyl-D-galactose was produced by substituting D-glucose with D-galactose. Based on the concentrations of D-glucose and Dgalactose as the starting materials, the yields were calculated to be 52 and 56%, respectively. These one-pot enzymatic approaches can be extended to include practical production of a variety of oligosaccharides by substituting nigerose phosphorylase with other β-D-glucose 1-phosphate-forming phosphorylases together with various carbohydrate acceptors.
Journal of Applied Glycoscience, 2011
Because numerous biologically functional enzymes generate Pi from labile phosphate esters such as... more Because numerous biologically functional enzymes generate Pi from labile phosphate esters such as ATP, pyrophosphate, or sugar 1-phosphates, the quantifi cation of Pi in the presence of these compounds is required for biochemical assays. The molybdate based colorimetry is one of the standard methods to quantify Pi. However, the method requires highly acidic conditions where the labile phosphate esters are often hydrolyzed. 1) To avoid the spontaneous hydrolyses, the colorimetric quantifi cation under weak acidic conditions is often used to measure Pi in the presence of labile biochemical phosphate esters, 1) such as monitoring the reaction catalyzed by phosphorylases. 2) However, the color development under these conditions is often affected by several compounds including phosphate esters, thus rendering accurate measurements diffi cult. Similarly, these methods cannot be used for continuous monitoring of enzymatic reactions, because molybdate inhibits these reactions. Several biosensors specifi c to Pi have been reported based on the measurement of the enzymatic consumption of oxygen detected with an oxygen electrode. The phosphatase-glucose oxidase sensor response to the concentration of Pi is based on the inhibition of the reaction catalyzed by the phosphatase that hydrolyzes glucose 6-phosphate into glucose, the substrate for glucose oxidase. 3,4) The purine nucleoside phosphorylase-xanthine oxidase sensor detects Pi through the oxidation of hypoxanthine produced by the phosphorolysis of inosine. 5 8) Oxygen-electrode based sensor with a tri
Journal of Applied Glycoscience, 2013
Bioscience of microbiota, food and health, 2012
We examined the effect of lacto-N-biose I (LNB) on Antigen (Ag)-specific responses of immune cell... more We examined the effect of lacto-N-biose I (LNB) on Antigen (Ag)-specific responses of immune cells. LNB exposure in vitro suppressed Ag-specific Interleukin (IL)-4 secretion of mouse splenocytes significantly. However, IL-4 secretion from CD4(+) T cells stimulated with anti-CD3ε did not changed significantly with LNB exposure. Additionally, Ag-specific Th1 cytokines did not change. Therefore LNB might suppress Ag-specific IL-4 through modification of Ag-presenting cells (APCs) in a manner independent of Th1-type immune development.
PloS one, 2014
We characterized Teth514_1788 and Teth514_1789, belonging to glycoside hydrolase family 130, from... more We characterized Teth514_1788 and Teth514_1789, belonging to glycoside hydrolase family 130, from Thermoanaerobacter sp. X-514. These two enzymes catalyzed the synthesis of 1,2-β-oligomannan using β-1,2-mannobiose and d-mannose as the optimal acceptors, respectively, in the presence of the donor α-d-mannose 1-phosphate. Kinetic analysis of the phosphorolytic reaction toward 1,2-β-oligomannan revealed that these enzymes followed a typical sequential Bi Bi mechanism. The kinetic parameters of the phosphorolysis of 1,2-β-oligomannan indicate that Teth514_1788 and Teth514_1789 prefer 1,2-β-oligomannans containing a DP ≥3 and β-1,2-Man2, respectively. These results indicate that the two enzymes are novel inverting phosphorylases that exhibit distinct chain-length specificities toward 1,2-β-oligomannan. Here, we propose 1,2-β-oligomannan:phosphate α-d-mannosyltransferase as the systematic name and 1,2-β-oligomannan phosphorylase as the short name for Teth514_1788 and β-1,2-mannobiose:phos...
Journal of Biological Chemistry, 2013
A gene cluster involved in N-glycan metabolism was identified in the genome of Bacteroides thetai... more A gene cluster involved in N-glycan metabolism was identified in the genome of Bacteroides thetaiotaomicron VPI-5482. This gene cluster encodes a major facilitator superfamily transporter, a starch utilization system-like transporter consisting of a TonB-dependent oligosaccharide transporter and an outer membrane lipoprotein, four glycoside hydrolases (α-mannosidase, β-N-acetylhexosaminidase, exo-α-sialidase, and endo-β-N-acetylglucosaminidase), and a phosphorylase (BT1033) with unknown function. It was demonstrated that BT1033 catalyzed the reversible phosphorolysis of β-1,4-d-mannosyl-N-acetyl-d-glucosamine in a typical sequential Bi Bi mechanism. These results indicate that BT1033 plays a crucial role as a key enzyme in the N-glycan catabolism where β-1,4-d-mannosyl-N-acetyl-d-glucosamine is liberated from N-glycans by sequential glycoside hydrolase-catalyzed reactions, transported into the cell, and intracellularly converted into α-d-mannose 1-phosphate and N-acetyl-d-glucosamin...
FEBS Letters, 2013
A novel phosphorylase was characterized as new member of glycoside hydrolase family 94 from the c... more A novel phosphorylase was characterized as new member of glycoside hydrolase family 94 from the cellulolytic bacterium Xanthomonas campestris and the fungus Neurospora crassa. The enzyme catalyzed reversible phosphorolysis of cellobionic acid. We propose 4‐O‐β‐d‐glucopyranosyl‐d‐gluconic acid: phosphate α‐d‐glucosyltransferase as the systematic name and cellobionic acid phosphorylase as the short names for the novel enzyme. Several cellulolytic fungi of the phylum Ascomycota also possess homologous proteins. We, therefore, suggest that the enzyme plays a crucial role in cellulose degradation where cellobionic acid as oxidized cellulolytic product is converted into α‐d‐glucose 1‐phosphate and d‐gluconic acid to enter glycolysis and the pentose phosphate pathway, respectively.
Bioscience, Biotechnology, and Biochemistry, 2014
4-O-β-d-Mannosyl-d-glucose phosphorylase (MGP), found in anaerobes, converts 4-O-β-d-mannosyl-d-g... more 4-O-β-d-Mannosyl-d-glucose phosphorylase (MGP), found in anaerobes, converts 4-O-β-d-mannosyl-d-glucose (Man-Glc) to α-d-mannosyl phosphate and d-glucose. It participates in mannan metabolism with cellobiose 2-epimerase (CE), which converts β-1,4-mannobiose to Man-Glc. A putative MGP gene is present in the genome of the thermophilic aerobe Rhodothermus marinus (Rm) upstream of the gene encoding CE. Konjac glucomannan enhanced production by R. marinus of MGP, CE, and extracellular mannan endo-1,4-β-mannosidase. Recombinant RmMGP catalyzed the phosphorolysis of Man-Glc through a sequential bi–bi mechanism involving ternary complex formation. Its molecular masses were 45 and 222 kDa under denaturing and nondenaturing conditions, respectively. Its pH and temperature optima were 6.5 and 75 °C, and it was stable between pH 5.5–8.3 and below 80 °C. In the reverse reaction, RmMGP had higher acceptor preferences for 6-deoxy-d-glucose and d-xylose than R. albus NE1 MGP. In contrast to R. albu...
Bioscience, Biotechnology, and Biochemistry, 2001
Bioscience, Biotechnology, and Biochemistry, 2007
cDNAs encoding three-glucosidases (HBGases I, II, and III) from European honeybees, Apis mellifer... more cDNAs encoding three-glucosidases (HBGases I, II, and III) from European honeybees, Apis mellifera, were cloned and sequenced, two of which were expressed in
Bioscience, Biotechnology, and Biochemistry, 2007
Two lacto-N-biose phosphorylase (LNBP) isozyme genes were cloned from Bifidobacterium bifidum JCM... more Two lacto-N-biose phosphorylase (LNBP) isozyme genes were cloned from Bifidobacterium bifidum JCM1254. Alignment of the amino acid sequences of LNBP and its homologs identified 24 completely conserved acidic amino acid residues. All single mutants of Bifidobacterium longum LNBP at residues other than D313N retained considerable activity, suggesting that Asp313 is the putative proton donor residue in LNBP.
Bioscience, Biotechnology, and Biochemistry, 2006
Bioscience, Biotechnology, and Biochemistry, 2012
Bioscience, Biotechnology, and Biochemistry, 2010
Bioscience, Biotechnology, and Biochemistry, 2009
FEBS letters, Jan 23, 2016
In Ruminococcus albus, either 4-O-β-d-mannosyl-d-glucose phosphorylase (RaMP1) and β-(1,4)-mannoo... more In Ruminococcus albus, either 4-O-β-d-mannosyl-d-glucose phosphorylase (RaMP1) and β-(1,4)-mannooligosaccharide phosphorylase (RaMP2) belong to two subfamilies of glycoside hydrolase family 130. The two enzymes phosphorylate β-mannosidic linkages at the non-reducing ends of their substrates, and have substantially diverse substrate specificity. The differences in their mechanism of substrate binding are yet not fully elucidated. Here, we report the crystal structures of RaMP1 with/without 4-O-β-d-mannosyl-d-glucose and RaMP2 with/without β-(1,4)-mannobiose. The structures of the two enzymes differ at the +1 subsite of the substrate binding pocket. Three loops are proposed to determine the different substrate specificities. Loop 3 interacts with the bound oligosaccharide. In RaMP1, His245 of loop 3 forms a hydrogen-bond network with the substrate through a water molecule, and is indispensible for substrate binding. This article is protected by copyright. All rights reserved.
Journal of Applied Glycoscience, 2014
Journal of Applied Glycoscience, 2001
Journal of Applied Glycoscience, 2002
Journal of Applied Glycoscience, 2011
Xylanase [EC 3.2.1.8] hydrolyzes the main chain of xylan (the major component of hemicelluloses),... more Xylanase [EC 3.2.1.8] hydrolyzes the main chain of xylan (the major component of hemicelluloses), consisting of β-1,4 xylosyl linkages in an endowise manner, and was also studied as an industrial enzyme for the utilization of unused biomass and a substitute for chemical bleach in paper manufacturing. 1 5) The natural substrate, xylan, is typically used for the characterization of xylanase. However, it is diffi cult to compare the characters of the enzymes determined with its natural substrate, xylan, for the following reasons: 1) Xylan is a heterogeneous polymer often heterogeneously decorated with α-L-arabinofuranosyl and/or 4-methyl-α-D-glucuronopyranosyl residues depending on its origin and lot; 6 11) 2) Several xylanases showed severe substrate inhibition of xylan. 12) Previously, we performed kinetic experiments of several xylanases using p-nitrophenyl-βxylobioside (XX-pNP) as the substrate. In such cases, no substrate inhibition was observed, and their activity could be evaluated accurately and reproducibly. 13) Xylanases belonging to the glycoside hydrolase family 10 (GH10) 14 16) have often been reported to hydrolyze cellobiosyl substrates such as p-nitrophenyl-β-cellobioside (GG-pNP). The ratio of the catalytic effi ciencies on XX-pNP and GG-pNP
Journal of Applied Glycoscience, 2014
One-pot enzymatic production of nigerose was demonstrated from abundantly available sugar resourc... more One-pot enzymatic production of nigerose was demonstrated from abundantly available sugar resources, including maltose, cellobiose, sucrose and starch. (i) 319 mM nigerose was generated from 500 mM maltose by the combined actions of maltose phosphorylase and nigerose phosphorylase, which share the same β-D-glucose 1-phosphate, in the presence of phosphate. The yield was 62% based on the concentration of maltose as the starting material. (ii) 129 mM nigerose was produced from 250 mM cellobiose by cellobiose phosphorylase and nigerose phosphorylase in the presence of phosphate, in combination with the enzymatic pathway to convert α-D-glucose 1-phosphate to β-D-glucose 1-phosphate via D-glucose 6-phosphate by the combined actions of α-phosphoglucomutase and β-phosphoglucomutase, resulting in a yield of 52%. (iii) 350 mM nigerose was produced from 500 mM sucrose by substituting cellobiose phosphorylase with sucrose phosphorylase and adding xylose isomerase, giving a yield of 67%. (iv) 270 mM nigerose was generated from 100 mg/mL starch and 500 mM D-glucose by the concomitant actions of glycogen phosphorylase, isoamylase, α-phosphoglucomutase, β-phosphoglucomutase and nigerose phosphorylase, in the presence of phosphate. In addition, 280 mM 3-O-α-D-glucopyranosyl-D-galactose was produced by substituting D-glucose with D-galactose. Based on the concentrations of D-glucose and Dgalactose as the starting materials, the yields were calculated to be 52 and 56%, respectively. These one-pot enzymatic approaches can be extended to include practical production of a variety of oligosaccharides by substituting nigerose phosphorylase with other β-D-glucose 1-phosphate-forming phosphorylases together with various carbohydrate acceptors.
Journal of Applied Glycoscience, 2011
Because numerous biologically functional enzymes generate Pi from labile phosphate esters such as... more Because numerous biologically functional enzymes generate Pi from labile phosphate esters such as ATP, pyrophosphate, or sugar 1-phosphates, the quantifi cation of Pi in the presence of these compounds is required for biochemical assays. The molybdate based colorimetry is one of the standard methods to quantify Pi. However, the method requires highly acidic conditions where the labile phosphate esters are often hydrolyzed. 1) To avoid the spontaneous hydrolyses, the colorimetric quantifi cation under weak acidic conditions is often used to measure Pi in the presence of labile biochemical phosphate esters, 1) such as monitoring the reaction catalyzed by phosphorylases. 2) However, the color development under these conditions is often affected by several compounds including phosphate esters, thus rendering accurate measurements diffi cult. Similarly, these methods cannot be used for continuous monitoring of enzymatic reactions, because molybdate inhibits these reactions. Several biosensors specifi c to Pi have been reported based on the measurement of the enzymatic consumption of oxygen detected with an oxygen electrode. The phosphatase-glucose oxidase sensor response to the concentration of Pi is based on the inhibition of the reaction catalyzed by the phosphatase that hydrolyzes glucose 6-phosphate into glucose, the substrate for glucose oxidase. 3,4) The purine nucleoside phosphorylase-xanthine oxidase sensor detects Pi through the oxidation of hypoxanthine produced by the phosphorolysis of inosine. 5 8) Oxygen-electrode based sensor with a tri
Journal of Applied Glycoscience, 2013
Bioscience of microbiota, food and health, 2012
We examined the effect of lacto-N-biose I (LNB) on Antigen (Ag)-specific responses of immune cell... more We examined the effect of lacto-N-biose I (LNB) on Antigen (Ag)-specific responses of immune cells. LNB exposure in vitro suppressed Ag-specific Interleukin (IL)-4 secretion of mouse splenocytes significantly. However, IL-4 secretion from CD4(+) T cells stimulated with anti-CD3ε did not changed significantly with LNB exposure. Additionally, Ag-specific Th1 cytokines did not change. Therefore LNB might suppress Ag-specific IL-4 through modification of Ag-presenting cells (APCs) in a manner independent of Th1-type immune development.
PloS one, 2014
We characterized Teth514_1788 and Teth514_1789, belonging to glycoside hydrolase family 130, from... more We characterized Teth514_1788 and Teth514_1789, belonging to glycoside hydrolase family 130, from Thermoanaerobacter sp. X-514. These two enzymes catalyzed the synthesis of 1,2-β-oligomannan using β-1,2-mannobiose and d-mannose as the optimal acceptors, respectively, in the presence of the donor α-d-mannose 1-phosphate. Kinetic analysis of the phosphorolytic reaction toward 1,2-β-oligomannan revealed that these enzymes followed a typical sequential Bi Bi mechanism. The kinetic parameters of the phosphorolysis of 1,2-β-oligomannan indicate that Teth514_1788 and Teth514_1789 prefer 1,2-β-oligomannans containing a DP ≥3 and β-1,2-Man2, respectively. These results indicate that the two enzymes are novel inverting phosphorylases that exhibit distinct chain-length specificities toward 1,2-β-oligomannan. Here, we propose 1,2-β-oligomannan:phosphate α-d-mannosyltransferase as the systematic name and 1,2-β-oligomannan phosphorylase as the short name for Teth514_1788 and β-1,2-mannobiose:phos...
Journal of Biological Chemistry, 2013
A gene cluster involved in N-glycan metabolism was identified in the genome of Bacteroides thetai... more A gene cluster involved in N-glycan metabolism was identified in the genome of Bacteroides thetaiotaomicron VPI-5482. This gene cluster encodes a major facilitator superfamily transporter, a starch utilization system-like transporter consisting of a TonB-dependent oligosaccharide transporter and an outer membrane lipoprotein, four glycoside hydrolases (α-mannosidase, β-N-acetylhexosaminidase, exo-α-sialidase, and endo-β-N-acetylglucosaminidase), and a phosphorylase (BT1033) with unknown function. It was demonstrated that BT1033 catalyzed the reversible phosphorolysis of β-1,4-d-mannosyl-N-acetyl-d-glucosamine in a typical sequential Bi Bi mechanism. These results indicate that BT1033 plays a crucial role as a key enzyme in the N-glycan catabolism where β-1,4-d-mannosyl-N-acetyl-d-glucosamine is liberated from N-glycans by sequential glycoside hydrolase-catalyzed reactions, transported into the cell, and intracellularly converted into α-d-mannose 1-phosphate and N-acetyl-d-glucosamin...
FEBS Letters, 2013
A novel phosphorylase was characterized as new member of glycoside hydrolase family 94 from the c... more A novel phosphorylase was characterized as new member of glycoside hydrolase family 94 from the cellulolytic bacterium Xanthomonas campestris and the fungus Neurospora crassa. The enzyme catalyzed reversible phosphorolysis of cellobionic acid. We propose 4‐O‐β‐d‐glucopyranosyl‐d‐gluconic acid: phosphate α‐d‐glucosyltransferase as the systematic name and cellobionic acid phosphorylase as the short names for the novel enzyme. Several cellulolytic fungi of the phylum Ascomycota also possess homologous proteins. We, therefore, suggest that the enzyme plays a crucial role in cellulose degradation where cellobionic acid as oxidized cellulolytic product is converted into α‐d‐glucose 1‐phosphate and d‐gluconic acid to enter glycolysis and the pentose phosphate pathway, respectively.
Bioscience, Biotechnology, and Biochemistry, 2014
4-O-β-d-Mannosyl-d-glucose phosphorylase (MGP), found in anaerobes, converts 4-O-β-d-mannosyl-d-g... more 4-O-β-d-Mannosyl-d-glucose phosphorylase (MGP), found in anaerobes, converts 4-O-β-d-mannosyl-d-glucose (Man-Glc) to α-d-mannosyl phosphate and d-glucose. It participates in mannan metabolism with cellobiose 2-epimerase (CE), which converts β-1,4-mannobiose to Man-Glc. A putative MGP gene is present in the genome of the thermophilic aerobe Rhodothermus marinus (Rm) upstream of the gene encoding CE. Konjac glucomannan enhanced production by R. marinus of MGP, CE, and extracellular mannan endo-1,4-β-mannosidase. Recombinant RmMGP catalyzed the phosphorolysis of Man-Glc through a sequential bi–bi mechanism involving ternary complex formation. Its molecular masses were 45 and 222 kDa under denaturing and nondenaturing conditions, respectively. Its pH and temperature optima were 6.5 and 75 °C, and it was stable between pH 5.5–8.3 and below 80 °C. In the reverse reaction, RmMGP had higher acceptor preferences for 6-deoxy-d-glucose and d-xylose than R. albus NE1 MGP. In contrast to R. albu...
Bioscience, Biotechnology, and Biochemistry, 2001
Bioscience, Biotechnology, and Biochemistry, 2007
cDNAs encoding three-glucosidases (HBGases I, II, and III) from European honeybees, Apis mellifer... more cDNAs encoding three-glucosidases (HBGases I, II, and III) from European honeybees, Apis mellifera, were cloned and sequenced, two of which were expressed in
Bioscience, Biotechnology, and Biochemistry, 2007
Two lacto-N-biose phosphorylase (LNBP) isozyme genes were cloned from Bifidobacterium bifidum JCM... more Two lacto-N-biose phosphorylase (LNBP) isozyme genes were cloned from Bifidobacterium bifidum JCM1254. Alignment of the amino acid sequences of LNBP and its homologs identified 24 completely conserved acidic amino acid residues. All single mutants of Bifidobacterium longum LNBP at residues other than D313N retained considerable activity, suggesting that Asp313 is the putative proton donor residue in LNBP.
Bioscience, Biotechnology, and Biochemistry, 2006
Bioscience, Biotechnology, and Biochemistry, 2012
Bioscience, Biotechnology, and Biochemistry, 2010
Bioscience, Biotechnology, and Biochemistry, 2009