The Experts below are selected from a list of 168 Experts worldwide ranked by ideXlab platform
Motomitsu Kitaoka - One of the best experts on this subject based on the ideXlab platform.
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Open–close structural change upon ligand binding and two magnesium ions required for the catalysis of N-Acetylhexosamine 1-kinase
Biochimica et biophysica acta, 2015Co-Authors: Mayo Sato, Mamoru Nishimoto, Motomitsu Kitaoka, Takatoshi Arakawa, Young-woo Nam, Shinya FushinobuAbstract:Infant gut-associated bifidobacteria possess a metabolic pathway to utilize lacto-N-biose (Gal-β1,3-GlcNAc) and galacto-N-biose (Gal-β1,3-GalNAc) from human milk and glycoconjugates specifically. In this pathway, N-Acetylhexosamine 1-kinase (NahK) catalyzes the phosphorylation of GlcNAc or GalNAc at the anomeric C1 position with ATP. Crystal structures of NahK have only been determined in the closed state. In this study, we determined open state structures of NahK in three different forms (apo, ADP complex, and ATP complex). A comparison of the open and closed state structures revealed an induced fit structural change defined by two rigid domains. ATP binds to the small N-terminal domain, and binding of the N-Acetylhexosamine substrate to the large C-terminal domain induces a closing conformational change with a rotation angle of 16°. In the nucleotide binding site, two magnesium ions bridging the α-γ and β-γ phosphates were identified. A mutational analysis indicated that a residue coordinating both of the two magnesium ions (Asp228) is essential for catalysis. The involvement of two magnesium ions in the catalytic machinery is structurally similar to the catalytic structures of protein kinases and aminoglycoside phosphotransferases, but distinct from the structures of other anomeric kinases or sugar 6-kinases. These findings help to elucidate the possible evolutionary adaptation of substrate specificities and induced fit mechanism.
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Identification of amino acid residues that determine the substrate preference of 1,3-β-galactosyl-N-Acetylhexosamine phosphorylase
Journal of Molecular Catalysis B: Enzymatic, 2012Co-Authors: Mamoru Nishimoto, Masahiro Nakajima, Shinya Fushinobu, Masafumi Hidaka, Motomitsu KitaokaAbstract:Abstract Three amino acid residues of 1,3-β-galactosyl- N -acetylhexosamine phosphorylase (GalHexNAcP) were assigned as the determinants of substrate preference for galacto- N -biose (GNB) and lacto- N -biose I (LNB) based on the three-dimensional structure of the protein. Mutants of GalHexNAcP from Bifidobacterium longum , which acts similarly on both GNB and LNB, were constructed and characterized. V162T mutation led to an increase in the selectivity on GNB. P161S and S336A mutations independently enhanced the selectivity on LNB. The alignment of amino acid sequences suggests that the activities of most homologous sequences are predictable by comparing the corresponding three residues.
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Substrate Specificity of N‐Acetylhexosamine Kinase Towards N‐Acetylgalactosamine Derivatives.
ChemInform, 2010Co-Authors: Li Cai, Motomitsu Kitaoka, Wanyi Guan, Jie Shen, Wenjun Wang, Wei Zhao, Crystal O’neil, Peng George WangAbstract:We report herein a bacterial N-Acetylhexosamine kinase, NahK, with broad substrate specificity towards structurally modified GalNAc analogues, and the production of a GalNAc-1-phosphate library using this kinase.
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A Chemoenzymatic Route to N‐Acetylglucosamine‐1‐phosphate Analogues: Substrate Specificity Investigations of N‐Acetylhexosamine 1‐Kinase.
ChemInform, 2009Co-Authors: Li Cai, Motomitsu Kitaoka, Wanyi Guan, Jie Shen, Chengfeng Xia, Wenlan Chen, Peng George WangAbstract:Reports an efficient chemoenzymatic production of an N-Acetylhexosamine 1-phophate analogues library by N-Acetylhexosamine 1-kinase (NahK) and describes the respective substrate specificity on this enzyme.
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substrate specificity of n acetylhexosamine kinase towards n acetylgalactosamine derivatives
Bioorganic & Medicinal Chemistry Letters, 2009Co-Authors: Motomitsu Kitaoka, Wanyi Guan, Jie Shen, Wenjun Wang, Wei Zhao, Crystal Oneil, Peng George WangAbstract:We report herein a bacterial N-Acetylhexosamine kinase, NahK, with broad substrate specificity towards structurally modified GalNAc analogues, and the production of a GalNAc-1-phosphate library using this kinase.
Peng George Wang - One of the best experts on this subject based on the ideXlab platform.
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efficient enzymatic synthesis of guanosine 5 diphosphate sugars and derivatives
Organic Letters, 2013Co-Authors: Yonghui Liu, Xi Chen, Peng George Wang, Wei Zhao, Yue WanAbstract:An N-Acetylhexosamine 1-kinase from Bifidobacterium infantis (NahK_15697), a guanosine 5′-diphosphate (GDP)-mannose pyrophosphorylase from Pyrococcus furiosus (PFManC), and an Escherichia coli inorganic pyrophosphatase (EcPpA) were used efficiently for a one-pot three-enzyme synthesis of GDP-mannose, GDP-glucose, their derivatives, and GDP-talose. This study represents the first facile and efficient enzymatic synthesis of GDP-sugars and derivatives starting from monosaccharides and derivatives.
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Substrate Promiscuity of N-Acetylhexosamine 1-Kinases
Molecules (Basel Switzerland), 2011Co-Authors: Yi Chen, Vireak Thon, Kam Lau, Li Cai, Hongzhi Cao, Vinod K. Tiwari, Peng George WangAbstract:N-Acetylhexosamine 1-kinase (NahK) catalyzes the direct addition of a phosphate from adenosine 5'-triphosphate (ATP) to the anomeric position of N-Acetylhexosamine and shows similar activity towards N-acetylglucosamine (GlcNAc) and N-acetylgalactosamine (GalNAc). Herein we report the cloning, characterization, and substrate specificity studies of two NahKs from Bifidobacterium infantis ATCC15697 and Bifidobacterium longum ATCC55813, respectively. A new capillary electrophoresis assay method has been developed for enzyme activity assays. Both enzymes have a good expression level in E. coli (180-185 mg/L culture) and can tolerate diverse modifications at C2 of GlcNAc and GalNAc. Various GlcNAc derivatives with C6, both C2 and C6, as well as both C2 and C3 modifications are tolerable substrates for the newly cloned NahKs. Quite interestingly, despite of their low activities toward glucose and galactose, the activities of both NahKs are much higher for mannose and some of its C2, C4, and C6 derivatives. These NahKs are excellent catalysts for enzymatic and chemoenzymatic synthesis of carbohydrates.
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Substrate Specificity of N‐Acetylhexosamine Kinase Towards N‐Acetylgalactosamine Derivatives.
ChemInform, 2010Co-Authors: Li Cai, Motomitsu Kitaoka, Wanyi Guan, Jie Shen, Wenjun Wang, Wei Zhao, Crystal O’neil, Peng George WangAbstract:We report herein a bacterial N-Acetylhexosamine kinase, NahK, with broad substrate specificity towards structurally modified GalNAc analogues, and the production of a GalNAc-1-phosphate library using this kinase.
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A Chemoenzymatic Route to N‐Acetylglucosamine‐1‐phosphate Analogues: Substrate Specificity Investigations of N‐Acetylhexosamine 1‐Kinase.
ChemInform, 2009Co-Authors: Li Cai, Motomitsu Kitaoka, Wanyi Guan, Jie Shen, Chengfeng Xia, Wenlan Chen, Peng George WangAbstract:Reports an efficient chemoenzymatic production of an N-Acetylhexosamine 1-phophate analogues library by N-Acetylhexosamine 1-kinase (NahK) and describes the respective substrate specificity on this enzyme.
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substrate specificity of n acetylhexosamine kinase towards n acetylgalactosamine derivatives
Bioorganic & Medicinal Chemistry Letters, 2009Co-Authors: Motomitsu Kitaoka, Wanyi Guan, Jie Shen, Wenjun Wang, Wei Zhao, Crystal Oneil, Peng George WangAbstract:We report herein a bacterial N-Acetylhexosamine kinase, NahK, with broad substrate specificity towards structurally modified GalNAc analogues, and the production of a GalNAc-1-phosphate library using this kinase.
Hayden Thompson - One of the best experts on this subject based on the ideXlab platform.
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Toward Closing the Gap between Hexoses and N-Acetlyhexosamines: Experimental and Computational Studies on the Collision-Induced Dissociation of Hexosamines.
The journal of physical chemistry. A, 2019Co-Authors: Cheng-chau Chiu, Hai Thi Huynh, Shang-ting Tsai, Hou-yu Lin, Po-jen Hsu, Huu Trong Phan, Arya Karumanthra, Hayden Thompson, Yu-chi Lee, Jer-lai KuoAbstract:Motivated by the fundamental difference in the reactivity of hexoses and N-Acetylhexosamines under collision-induced dissociation (CID) mass spectrometry conditions, we have investigated the CID of...
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toward closing the gap between hexoses and n acetlyhexosamines experimental and computational studies on the collision induced dissociation of hexosamines
Journal of Physical Chemistry A, 2019Co-Authors: Cheng-chau Chiu, Hai Thi Huynh, Shang-ting Tsai, Huu Trong Phan, Arya Karumanthra, Hayden ThompsonAbstract:Motivated by the fundamental difference in the reactivity of hexoses and N-Acetylhexosamines under collision-induced dissociation (CID) mass spectrometry conditions, we have investigated the CID of two hexosamines, glucosamine (GlcN) and galactosamine (GalN), experimentally and computationally. Both hexosamines undergo ring-opening and then dissociate via the 0,2A and the 0,3A (0,3X) cross-ring cleavage channels. The preference for the ring-opening is similar to the behavior of N-Acetylhexosamines and explains, why the two anomers of the same sugar give the same mass spectrum. While the spectrum for GlcN is dominated by the 0,2A signal, the signal intensities for both 0,2A and the 0,3A (0,3X) dissociation channels are comparable for GalN, which allows GlcN and GalN to be distinguished easily. Calculations at MP2 level of theory indicate that this is related to the differences in the relative barrier heights for the 0,2A and the 0,3A (0,3X) cross-ring cleavage channels. This, in return, reflects the circum...
Mamoru Nishimoto - One of the best experts on this subject based on the ideXlab platform.
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Open–close structural change upon ligand binding and two magnesium ions required for the catalysis of N-Acetylhexosamine 1-kinase
Biochimica et biophysica acta, 2015Co-Authors: Mayo Sato, Mamoru Nishimoto, Motomitsu Kitaoka, Takatoshi Arakawa, Young-woo Nam, Shinya FushinobuAbstract:Infant gut-associated bifidobacteria possess a metabolic pathway to utilize lacto-N-biose (Gal-β1,3-GlcNAc) and galacto-N-biose (Gal-β1,3-GalNAc) from human milk and glycoconjugates specifically. In this pathway, N-Acetylhexosamine 1-kinase (NahK) catalyzes the phosphorylation of GlcNAc or GalNAc at the anomeric C1 position with ATP. Crystal structures of NahK have only been determined in the closed state. In this study, we determined open state structures of NahK in three different forms (apo, ADP complex, and ATP complex). A comparison of the open and closed state structures revealed an induced fit structural change defined by two rigid domains. ATP binds to the small N-terminal domain, and binding of the N-Acetylhexosamine substrate to the large C-terminal domain induces a closing conformational change with a rotation angle of 16°. In the nucleotide binding site, two magnesium ions bridging the α-γ and β-γ phosphates were identified. A mutational analysis indicated that a residue coordinating both of the two magnesium ions (Asp228) is essential for catalysis. The involvement of two magnesium ions in the catalytic machinery is structurally similar to the catalytic structures of protein kinases and aminoglycoside phosphotransferases, but distinct from the structures of other anomeric kinases or sugar 6-kinases. These findings help to elucidate the possible evolutionary adaptation of substrate specificities and induced fit mechanism.
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Identification of amino acid residues that determine the substrate preference of 1,3-β-galactosyl-N-Acetylhexosamine phosphorylase
Journal of Molecular Catalysis B: Enzymatic, 2012Co-Authors: Mamoru Nishimoto, Masahiro Nakajima, Shinya Fushinobu, Masafumi Hidaka, Motomitsu KitaokaAbstract:Abstract Three amino acid residues of 1,3-β-galactosyl- N -acetylhexosamine phosphorylase (GalHexNAcP) were assigned as the determinants of substrate preference for galacto- N -biose (GNB) and lacto- N -biose I (LNB) based on the three-dimensional structure of the protein. Mutants of GalHexNAcP from Bifidobacterium longum , which acts similarly on both GNB and LNB, were constructed and characterized. V162T mutation led to an increase in the selectivity on GNB. P161S and S336A mutations independently enhanced the selectivity on LNB. The alignment of amino acid sequences suggests that the activities of most homologous sequences are predictable by comparing the corresponding three residues.
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Characterization of β-1,3-galactosyl-N-Acetylhexosamine phosphorylase from Propionibacterium acnes
Applied Microbiology and Biotechnology, 2009Co-Authors: Masahiro Nakajima, Mamoru Nishimoto, Motomitsu KitaokaAbstract:Homologs of the β-1,3-galactosyl- N -acetylhexosamine phosphorylase (GalHexNAcP) gene ( gnpA ) were cloned from the genomic DNA of Propionibacterium acnes JCM6425 and P. acnes JCM6473, showing 99.9% and 97.9% nucleotide sequence identity, respectively, with the ppa0083 gene from the genome-sequenced P. acnes KPA171202. No gnpA gene was detected in the genomic DNA of type strain P. acnes ATCC25746. The recombinant enzyme from P. acnes JCM6425 (GnpA) showed approximately 70 times higher specific activity of phosphorolysis on galacto- N -biose (Galβ1→3GalNAc, GNB) than that on lacto- N -biose I (Galβ1→3GlcNAc). K _m value for GnpA on GNB was high, but GnpA did not exhibit activity on any derivatives of GNB examined. These results indicate that GnpA is GalHexNAcP which should be classified as galacto- N -biose phosphorylase. The large k _cat value of GnpA on GalNAc suggests that GnpA would be a useful catalyst for the synthesis of GNB.
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identification of n acetylhexosamine 1 kinase in the complete lacto n biose i galacto n biose metabolic pathway in bifidobacterium longum
Applied and Environmental Microbiology, 2007Co-Authors: Mamoru Nishimoto, Motomitsu KitaokaAbstract:We have determined the functions of the enzymes encoded by the lnpB, lnpC, and lnpD genes, located downstream of the lacto-N-biose phosphorylase gene (lnpA), in Bifidobacterium longum JCM1217. The lnpB gene encodes a novel kinase, N-Acetylhexosamine 1-kinase, which produces N-Acetylhexosamine 1-phosphate; the lnpC gene encodes UDP-glucose hexose 1-phosphate uridylyltransferase, which is also active on N-Acetylhexosamine 1-phosphate; and the lnpD gene encodes a UDP-glucose 4-epimerase, which is active on both UDP-galactose and UDP-N-acetylgalactosamine. These results suggest that the gene operon lnpABCD encodes a previously undescribed lacto-N-biose I/galacto-N-biose metabolic pathway that is involved in the intestinal colonization of bifidobacteria and that utilizes lacto-N-biose I from human milk oligosaccharides or galacto-N-biose from mucin sugars.
Wanyi Guan - One of the best experts on this subject based on the ideXlab platform.
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Active sites of N-Acetylhexosamine 1-kinase from Bifidobacterium longum
Wei sheng wu xue bao = Acta microbiologica Sinica, 2016Co-Authors: Wanyi Guan, Jing Bai, Tianhui Zhou, Baohua ZhaoAbstract:OBJECTIVE To study the active sites of N-Acetylhexosamine 1-kinase (NahK) from Bifidobacterium longum JCM12 17. METHODS We obtained expression strains of 10 single-mutants at 4 sites of NahK by site-directed mutagenesis, and expressed and purified both wild-type (WT) and mutant enzymes. Then, their optimum pH and optimum concentration of Mg²⁺ were determined by DNS assay and NADH-coupled microplate photometric assay, and their kinetic parameters were measured. RESULTS Four mutants (D208A, D208N, D208E and I24A) lost most part of the catalytic activity. The optimum pH of mutants H31A, H31V, F247A and I24V switched from pH 7.5 (for WT) to pH 7.0, and the optimum concentration of Mg²⁺ of mutants H31A and F247A increased to 10 mmol/L from 5 mmol/L (for WT). The kinetic parameters of WT and mutants indicate that mutant F247Y had higher enzymatic activity toward GlcNAc, GalNAc and ATP than WT. CONCLUSION The key amino acids that affect the catalytic activity of NahK were determined by site-directed mutagenesis, and together with the mutant that has higher catalytic efficiency, this has laid a foundation for further modification and evolution of NahK.
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Substrate Specificity of N‐Acetylhexosamine Kinase Towards N‐Acetylgalactosamine Derivatives.
ChemInform, 2010Co-Authors: Li Cai, Motomitsu Kitaoka, Wanyi Guan, Jie Shen, Wenjun Wang, Wei Zhao, Crystal O’neil, Peng George WangAbstract:We report herein a bacterial N-Acetylhexosamine kinase, NahK, with broad substrate specificity towards structurally modified GalNAc analogues, and the production of a GalNAc-1-phosphate library using this kinase.
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A Chemoenzymatic Route to N‐Acetylglucosamine‐1‐phosphate Analogues: Substrate Specificity Investigations of N‐Acetylhexosamine 1‐Kinase.
ChemInform, 2009Co-Authors: Li Cai, Motomitsu Kitaoka, Wanyi Guan, Jie Shen, Chengfeng Xia, Wenlan Chen, Peng George WangAbstract:Reports an efficient chemoenzymatic production of an N-Acetylhexosamine 1-phophate analogues library by N-Acetylhexosamine 1-kinase (NahK) and describes the respective substrate specificity on this enzyme.
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substrate specificity of n acetylhexosamine kinase towards n acetylgalactosamine derivatives
Bioorganic & Medicinal Chemistry Letters, 2009Co-Authors: Motomitsu Kitaoka, Wanyi Guan, Jie Shen, Wenjun Wang, Wei Zhao, Crystal Oneil, Peng George WangAbstract:We report herein a bacterial N-Acetylhexosamine kinase, NahK, with broad substrate specificity towards structurally modified GalNAc analogues, and the production of a GalNAc-1-phosphate library using this kinase.
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A chemoenzymatic route to N-acetylglucosamine-1-phosphate analogues: substrate specificity investigations of N-Acetylhexosamine 1-kinase
Chemical communications (Cambridge England), 2009Co-Authors: Li Cai, Motomitsu Kitaoka, Wanyi Guan, Jie Shen, Chengfeng Xia, Wenlan Chen, Peng George WangAbstract:Reports an efficient chemoenzymatic production of an N-Acetylhexosamine 1-phophate analogues library by N-Acetylhexosamine 1-kinase (NahK) and describes the respective substrate specificity on this enzyme.
