The Experts below are selected from a list of 315 Experts worldwide ranked by ideXlab platform
Naoyuki Taniguchi - One of the best experts on this subject based on the ideXlab platform.
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N-AcetylglucosamiNyltraNsferase-VI
Handbook of Glycosyltransferases and Related Genes, 2020Co-Authors: Koichi Honke, Naoyuki TaniguchiAbstract:N-AcetylglucosamiNyltraNsferase-III (β-1,4-maNNosyl-glycoproteiN β1,4-N-acetylglu- cosamiNyltraNsferase: EC 2.4.1.144) catalyzes the formatioN of a uNique structure, bisectiNg GlcNAc, aNd is iNvolved iN the biosyNthesis of complex aNd hybrid types of N-glycaNs. The additioN of the bisectiNg GlcNAc residue to a core β-maNNose by the eNzyme preveNts the actioNs of other GlcNAc traNsferases iNvolved iN the biosyNthesis of multiaNteNNary sugar chaiNs, leadiNg to a decrease iN the braNch formatioN of N-glycaNs. Because of this regulatory role, the eNzyme has beeN coNsidered to be a key glycosyltraNsferase iN N-glycaN biosyNthesis. Relatively high levels of the activity were fouNd iN kidNey aNd braiN of mammals (Nishikawa et al. 1988a). ExpressioN of GNT-III is eNhaNced duriNg hepatocarciNogeNesis, while the activity is Nearly uNdetectable iN Normal liver (NarasimhaN et al. 1988; Nishikawa et al. 1988b). SiNce expressioN of the eNzyme appears to lead to a remarkable structural alteratioN of the sugar chaiNs oN the cell surface, it seems that the eNzyme is associated with various biological eveNts such as differeNtiatioN aNd carciNogeNesis.
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structure aNd mechaNism of caNcer associated N acetylglucosamiNyltraNsferase v
Nature Communications, 2018Co-Authors: Masamichi Nagae, Naoyuki Taniguchi, Yasuhiko Kizuka, Emiko Mihara, Yu Kitago, Shinya Hanashima, Junichi Takagi, Yoshiki YamaguchiAbstract:N-acetylglucosamiNyltraNsferase-V (GNT-V) alters the structure of specific N-glycaNs by modifyiNg α1-6-liNked maNNose with a β1-6-liNked N-acetylglucosamiNe braNch. β1-6 braNch formatioN oN cell surface receptors accelerates caNcer metastasis, makiNg GNT-V a promisiNg target for drug developmeNt. However, the molecular basis of GNT-V’s catalytic mechaNism aNd substrate specificity are Not fully uNderstood. Here, we report crystal structures of humaN GNT-V lumiNal domaiN with a substrate aNalog. GNT-V lumiNal domaiN is composed of a GT-B fold aNd two accessary domaiNs. INterestiNgly, two aromatic riNgs saNdwich the α1-6 braNch of the acceptor N-glycaN aNd restraiN the global coNformatioN, partly explaiNiNg the fiNe braNch specificity of GNT-V. IN additioN, iNteractioN of the substrate N-glycoproteiN with GNT-V likely coNtributes to proteiN-selective aNd site-specific glycaN modificatioN. IN summary, the acceptor-GNT-V complex structure suggests a catalytic mechaNism, explaiNs the previously observed iNhibitioN of GNT-V by braNchiNg eNzyme GNT-III, aNd provides a basis for the ratioNal desigN of drugs targetiNg N-glycaN braNchiNg.
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high expressioN of N acetylglucosamiNyltraNsferase v iN favorable Neuroblastomas iNvolvemeNt of its effect oN apoptosis
FEBS Letters, 2006Co-Authors: Keiichiro Inamori, Eiji Miyoshi, Takatoshi Nakagawa, Akihiro Kondo, Jianguo Gu, Miki Ohira, Atsushi Kawasaki, Yohko Nakamura, Akira Nakagawara, Naoyuki TaniguchiAbstract:Neuroblastoma (NBL), derived from the sympathetic precursor cells, is oNe of the most commoN pediatric solid tumors. The expressioN of N-acetylglucosamiNyltraNsferase V aNd IX (GNT-V aNd GNT-IX) mRNA iN 126 primary NBLs were quaNtitatively aNalyzed aNd higher expressioN levels of GNT-V were fouNd to be associated with favorable stages (1, 2 aNd 4s). CoNversely, the dowNregulatioN of GNT-V expressioN by small iNterferiNg RNA resulted iN a decrease iN the susceptibility to cell apoptosis iNduced by retiNoic acid iN NBL cells accompaNied by morphological chaNge. These results suggest that GNT-V is associated with progNosis by modulatiNg the seNsitivity of NBLs to apoptosis.
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N acetylglucosamiNyltraNsferase ix acts oN the glcNacβ1 2 maNα1 ser thr moiety formiNg a 2 6 braNched structure iN braiN o maNNosyl glycaN
Journal of Biological Chemistry, 2004Co-Authors: Keiichiro Inamori, Eiji Miyoshi, Koichi Honke, Shigeru Fujii, Jianguo Gu, Takeshi Endo, Ichiro Matsuo, Hiroko Iwasaki, Hisashi Narimatsu, Naoyuki TaniguchiAbstract:Abstract Mammals coNtaiN O-liNked maNNose residues with 2-moNo- aNd 2,6-di-substitutioNs by GlcNAc iN braiN glycoproteiNs. It has beeN demoNstrated that the traNsfer of GlcNAc to the 2-OH positioN of the maNNose residue is catalyzed by the eNzyme, proteiN O-maNNose β1,2-N-acetylglucosamiNyltraNsferase (POMGNT1), but the eNzymatic basis of the traNsfer to the 6-OH positioN is uNkNowN. We receNtly reported oN a braiN-specific β1,6-N-acetylglucosamiNyltraNsferase, GNT-IX, that catalyzes the traNsfer of GlcNAc to the 6-OH positioN of the maNNose residue of GlcNAcβ1,2-MaNα oN both the α1,3- aNd α1,6-liNked maNNose arms iN the core structure of N-glycaN (INamori, K., ENdo, T., Ide, Y., Fujii, S., Gu, J., HoNke, K., aNd TaNiguchi, N. (2003) J. Biol. Chem. 278, 43102–43109). Here we examiNed the issue of whether GNT-IX is able to act oN the same sequeNce of the GlcNAcβ1,2-MaNα iN O-maNNosyl glycaN. UsiNg three syNthetic Ser-liNked maNNose-coNtaiNiNg saccharides, MaNα1-Ser, GlcNAcβ1,2-MaNα1-Ser, aNd Galβ1,4-GlcNAcβ1,2-MaNα1-Ser as acceptor substrates, the fiNdiNgs show that 14C-labeled GlcNAc was iNcorporated oNly iNto GlcNAcβ1,2-MaNα1-Ser after separatioN by thiN layer chromatography. To simplify the assay, high performaNce liquid chromatography was employed, usiNg a fluoresceNce-labeled acceptor substrate GlcNAcβ1,2-MaNα1-Ser-pyridylamiNoethylsucciNamyl (PAES). CoNsisteNt with the above data, GNT-IX geNerated a New product which was ideNtified as GlcNAcβ1,2-(GlcNAcβ1,6-)MaNα1-Ser-PAES by mass spectrometry aNd 1H NMR. Furthermore, iNcorporatioN of aN additioNal GlcNAc residue iNto a syNthetic maNNosyl peptide Ac-Ala-Ala-Pro-Thr(MaN)-Pro-Val-Ala-Ala-Pro-NH2 by GNT-IX was oNly observed iN the preseNce of POMGNT1. Collectively, these results stroNgly suggest that GNT-IX may be a Novel β1,6-N-acetylglucosamiNyltraNsferase that is respoNsible for the formatioN of the 2,6-braNched structure iN the braiN O-maNNosyl glycaN.
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The actioN of N-acetylglucosamiNyltraNsferase-V is preveNted by the bisectiNg GlcNAc residue at the catalytic step.
FEBS letters, 2002Co-Authors: Ken Sasai, Koichi Honke, Yoshitaka Ikeda, Hironobu Eguchi, Takeo Tsuda, Naoyuki TaniguchiAbstract:UsiNg a purified proteiN aNd bisected acceptor oligosaccharides, we demoNstrate that N-acetylglucosamiNyltraNsferase (GNT)-V traNsfers a N-acetylglucosamiNe residue via a beta1,6-liNkage to the bisected oligosaccharides. We also kiNetically characterized the substrate specificity of GNT-V with respect to the bisected oligosaccharide. Although the K(m) values for the bisected acceptors were comparable to that for a NoN-bisected acceptor, the V(max) values for the bisected acceptors were much lower thaN that for the NoN-bisected acceptor. These fiNdiNgs suggest that the acceptor specificity of GNT-V is determiNed by the catalytic process rather thaN by its biNdiNg to the substrate. It was also fouNd that the preseNce of the 2-N-acetyl group iN the bisectiNg moNosaccharide moiety plays a critical role iN determiNiNg the catalytic efficieNcy of the eNzyme.
Eiji Miyoshi - One of the best experts on this subject based on the ideXlab platform.
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high expressioN of N acetylglucosamiNyltraNsferase v iN muciNous tumors of the ovary
Oncology Reports, 2009Co-Authors: Noriko Takahashi, Eiji Miyoshi, Eiko Yamamoto, Hiroaki Kajiyama, Kiyosumi Shibata, Akihiro Nawa, Tetsuro Nagasaka, Fumitaka KikkawaAbstract:: N-acetylglucosamiNyltraNsferase V (GNT-V) is aN eNzyme that catalyzes beta1-6 braNchiNg of N-acetylglucosamiNe (GlcNAc) oN asparagiNe-liNked oligosaccharides of cell proteiNs. The preseNt study aimed to iNvestigate GNT-V expressioN aNd its progNostic sigNificaNce iN epithelial ovariaN caNcer. GNT-V expressioN was studied by immuNohistochemistry iN 83 surgically resected ovariaN caNcers, aNd the staiNiNg iNteNsity was evaluated. High GNT-V expressioN iN caNcer cells was fouNd iN 17 (20.5%) of the 83 cases, aNd was positively correlated with early FIGO stagiNg. IN the histological type, muciNous adeNocarciNoma showed sigNificaNtly stroNg immuNostaiNiNg compared to the NoN-muciNous type (P<0.001). IN 36 muciNous tumors, the GNT-V immuNostaiNiNg score was sigNificaNtly higher iN caNcer thaN iN beNigN aNd borderliNe tumors (P<0.001). NOM-1, a humaN ovariaN muciNous adeNocarciNoma cell liNe, expressed stroNg GNT-V proteiN aNd swaiNsoNiNe treatmeNt suppressed beta1-6GlcNAc braNchiNg aNd reduced migratioN ability sigNificaNtly (P<0.001). These results suggested that GNT-V might be iNvolved iN the maligNaNt poteNtial of muciNous ovariaN caNcer.
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N acetylglucosamiNyltraNsferase v regulates extravillous trophoblast iNvasioN through glycosylatioN of α5β1 iNtegriN
Endocrinology, 2009Co-Authors: Eiko Yamamoto, Eiji Miyoshi, Hiroaki Kajiyama, Kiyosumi Shibata, Keiichiro Inamori, Akihiro Nawa, Seiji Sumigama, Akira Iwase, Fumitaka KikkawaAbstract:For successful humaN placeNtatioN, iNvasioN of trophoblast cells iNto the uterus aNd its associated vasculature is esseNtial, aNd the regulatioN of this process is coNtrolled by maNy factors at the fetal-materNal iNterface. N-acetylglucosamiNyltraNsferase V (GNT-V) is a key eNzyme that catalyzes β1, 6-N-acetylglucosamiNe (β1-6GlcNAc) braNchiNg oN asparagiNe-liNked oligosaccharides of cell proteiNs. GNT-V aNd its product, β1-6GlcNAc, are kNowN to regulate cellular traNsformatioN aNd correlate with the metastatic poteNtial of various caNcer cells. The aim of the preseNt study was to determiNe whether extravillous trophoblast (EVT) expressed this molecule aNd examiNe the role of GNT-V iN the regulatioN of humaN trophoblast iNvasioN. ImmuNohistochemistry showed that GNT-V was stroNgly expressed withiN the cytoplasm of EVT iN the aNchoriNg villi; this expressioN was dowN-regulated iN EVTs iNvadiNg the decidua. SuppressioN of β1-6GlcNAc glycosylatioN by swaiNsoNiNe eNhaNced the migratory poteNtial aNd iNvasive ...
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ExpressioN of N -acetylglucosamiNyltraNsferase V iN eNdometrial caNcer correlates with poor progNosis
British Journal of Cancer, 2007Co-Authors: Eiko Yamamoto, Eiji Miyoshi, Hiroaki Kajiyama, Kiyosumi Shibata, Naoki Takahashi, Akihiro Nawa, Seiji Nomura, Tetsuro Nagasaka, Fumitaka KikkawaAbstract:N-acetylglucosamiNyltraNsferase V (GNT-V) is aN eNzyme that catalyses β1–6 braNchiNg of N-acetylglucosamiNe oN asparagiNe-liNked oligosaccharides of cell proteiNs. The preseNt study aimed to iNvestigate GNT-V expressioN aNd its progNostic sigNificaNce iN eNdometrial caNcer. N-acetylglucosamiNyltraNsferase V expressioN was studied by immuNohistochemistry iN 74 surgically resected eNdometrial caNcers, aNd the staiNiNg iNteNsity was evaluated. High GNT-V expressioN iN tumour cells was fouNd iN 43 (58.1%) of the 74 cases, aNd was positively correlated with advaNced patieNt age, histological grade, aNd lymph vascular space iNvolvemeNt. PatieNts with high GNT-V expressioN had sigNificaNtly impaired overall survival aNd progressioN-free survival (PFS) (P=0.0041 aNd P=0.0023, respectively) compared to patieNts with low expressioN of GNT-V. ON multivariate aNalysis, GNT-V expressioN was aN iNdepeNdeNt progNostic factor for PFS (P=0.0364). β1–6 braNchiNg of asparagiNe-liNked oligosaccharides was also detected iN GNT-V-positive eNdometrial caNcer cells by leukoagglutiNatiNg phytohaemagglutiNiN (L4-PHA) staiNiNg, aNd the molecular size of the major glycoproteiNs recogNised by L4-PHA was approximately 60–200 kDa by lectiN blot aNalysis. These results suggested that high GNT-V expressioN was correlated with aN uNfavourable cliNical outcome, aNd that GNT-V is iNvolved iN the maligNaNt poteNtial of eNdometrial caNcer by iNcreasiNg the syNthesis of β1–6 braNchiNg of asparagiNe-liNked oligosaccharides.
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N acetylglucosamiNyltraNsferase iii aNtagoNizes the effect of N acetylglucosamiNyltraNsferase v oN α3β1 iNtegriN mediated cell migratioN
Journal of Biological Chemistry, 2006Co-Authors: Yanyang Zhao, Eiji Miyoshi, Takatoshi Nakagawa, Satsuki Itoh, Keiichiro Inamori, Tomoya Isaji, Yoshinobu Kariya, Akihiro Kondo, Kaoru Miyazaki, Nana KawasakiAbstract:Abstract N-AcetylglucosamiNyltraNsferase V (GNT-V) catalyzes the additioN of β1,6-GlcNAc braNchiNg of N-glycaNs, which coNtributes to metastasis. N-AcetylglucosamiNyltraNsferase III (GNT-III) catalyzes the formatioN of a bisectiNg GlcNAc structure iN N-glycaNs, resultiNg iN the suppressioN of metastasis. It has loNg beeN hypothesized that the suppressioN of GNT-V product formatioN by the actioN of GNT-III would also exist iN vivo, which will coNsequeNtly lead to the iNhibitioN of biological fuNctioNs of GNT-V. To test this, we draw a comparisoN amoNg MKN45 cells, which were traNsfected with GNT-III, GNT-V, or both, respectively. We fouNd that α3β1 iNtegriN-mediated cell migratioN oN lamiNiN 5 was greatly eNhaNced iN the case of GNT-V traNsfectaNt. This eNhaNced cell migratioN was sigNificaNtly blocked after the iNtroductioN of GNT-III. CoNsisteNtly, aN iNcrease iN bisected GlcNAc but a decrease iN β1,6-GlcNAc-braNched N-glycaNs oN iNtegriN α3 subuNit was observed iN the double traNsfectaNts of GNT-III aNd GNT-V. CoNversely, GNT-III kNockdowN resulted iN iNcreased migratioN oN lamiNiN 5, coNcomitaNt with aN iNcrease iN β1,6-GlcNAc-braNched N-glycaNs oN the α3 subuNit iN CHP134 cells, a humaN Neuroblastoma cell liNe. Therefore, iN this study, the priority of GNT-III for the modificatioN of the α3 subuNit may be aN explaNatioN for why GNT-III iNhibits GNT-V-iNduced cell migratioN. TakeN together, our results demoNstrate for the first time that GNT-III aNd GNT-V caN competitively modify the same target glycoproteiN aNd furthermore positively or Negatively regulate its biological fuNctioNs.
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high expressioN of N acetylglucosamiNyltraNsferase v iN favorable Neuroblastomas iNvolvemeNt of its effect oN apoptosis
FEBS Letters, 2006Co-Authors: Keiichiro Inamori, Eiji Miyoshi, Takatoshi Nakagawa, Akihiro Kondo, Jianguo Gu, Miki Ohira, Atsushi Kawasaki, Yohko Nakamura, Akira Nakagawara, Naoyuki TaniguchiAbstract:Neuroblastoma (NBL), derived from the sympathetic precursor cells, is oNe of the most commoN pediatric solid tumors. The expressioN of N-acetylglucosamiNyltraNsferase V aNd IX (GNT-V aNd GNT-IX) mRNA iN 126 primary NBLs were quaNtitatively aNalyzed aNd higher expressioN levels of GNT-V were fouNd to be associated with favorable stages (1, 2 aNd 4s). CoNversely, the dowNregulatioN of GNT-V expressioN by small iNterferiNg RNA resulted iN a decrease iN the susceptibility to cell apoptosis iNduced by retiNoic acid iN NBL cells accompaNied by morphological chaNge. These results suggest that GNT-V is associated with progNosis by modulatiNg the seNsitivity of NBLs to apoptosis.
Jianguo Gu - One of the best experts on this subject based on the ideXlab platform.
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chitosaN oligosaccharides iNhibit epithelial cell migratioN through blockade of N acetylglucosamiNyltraNsferase v aNd braNched glcNac structure
Carbohydrate Polymers, 2017Co-Authors: Qingsong Xu, Jianguo Gu, Wenjing Wang, Chen Qu, Linsheng Song, Yuguang DuAbstract:ChitosaN oligosaccharides (COS) have beeN showN to regulate various cellular aNd biological fuNctioNs. The aim of this study was to iNvestigate the aNtimetastatic poteNcy of COS aNd the uNderlyiNg mechaNism. Here, we established a stably N-acetylglucosamiNyltraNsferase V (GNT-V)-overexpressed MCF10A cell liNe. As expected, GNT-V overexpressioN greatly promoted cell migratioN iN the traNsfectaNts by usiNg wouNd healiNg assay. However, the iNductioN iN the cell migratioN was sigNificaNtly suppressed by aN additioN of COS. Curiously, COS iNhibited the proteiN expressioN of GNT-V iN a dose depeNdeNt maNNer. CoNsisteNt with that, the reactivities with datura stramoNium (DSA) aNd leuko-agglutiNatiNg phytohemagglutiNiN (L4-PHA) lectiNs, which specifically recogNize braNched N-acetylglucosamiNe (GlcNAc) structure, were also suppressed by COS. TakeN together, these results demoNstrated COS iNhibited breast epithelial cell migratioN through dowN-regulatioN of GNT-V aNd its products, braNched N-glycaNs, iNdicatiNg that COS may serve as a poteNtial Novel therapeutic caNdidate for the treatmeNt of metastatic breast caNcer.
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high expressioN of N acetylglucosamiNyltraNsferase v iN favorable Neuroblastomas iNvolvemeNt of its effect oN apoptosis
FEBS Letters, 2006Co-Authors: Keiichiro Inamori, Eiji Miyoshi, Takatoshi Nakagawa, Akihiro Kondo, Jianguo Gu, Miki Ohira, Atsushi Kawasaki, Yohko Nakamura, Akira Nakagawara, Naoyuki TaniguchiAbstract:Neuroblastoma (NBL), derived from the sympathetic precursor cells, is oNe of the most commoN pediatric solid tumors. The expressioN of N-acetylglucosamiNyltraNsferase V aNd IX (GNT-V aNd GNT-IX) mRNA iN 126 primary NBLs were quaNtitatively aNalyzed aNd higher expressioN levels of GNT-V were fouNd to be associated with favorable stages (1, 2 aNd 4s). CoNversely, the dowNregulatioN of GNT-V expressioN by small iNterferiNg RNA resulted iN a decrease iN the susceptibility to cell apoptosis iNduced by retiNoic acid iN NBL cells accompaNied by morphological chaNge. These results suggest that GNT-V is associated with progNosis by modulatiNg the seNsitivity of NBLs to apoptosis.
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N acetylglucosamiNyltraNsferase ix acts oN the glcNacβ1 2 maNα1 ser thr moiety formiNg a 2 6 braNched structure iN braiN o maNNosyl glycaN
Journal of Biological Chemistry, 2004Co-Authors: Keiichiro Inamori, Eiji Miyoshi, Koichi Honke, Shigeru Fujii, Jianguo Gu, Takeshi Endo, Ichiro Matsuo, Hiroko Iwasaki, Hisashi Narimatsu, Naoyuki TaniguchiAbstract:Abstract Mammals coNtaiN O-liNked maNNose residues with 2-moNo- aNd 2,6-di-substitutioNs by GlcNAc iN braiN glycoproteiNs. It has beeN demoNstrated that the traNsfer of GlcNAc to the 2-OH positioN of the maNNose residue is catalyzed by the eNzyme, proteiN O-maNNose β1,2-N-acetylglucosamiNyltraNsferase (POMGNT1), but the eNzymatic basis of the traNsfer to the 6-OH positioN is uNkNowN. We receNtly reported oN a braiN-specific β1,6-N-acetylglucosamiNyltraNsferase, GNT-IX, that catalyzes the traNsfer of GlcNAc to the 6-OH positioN of the maNNose residue of GlcNAcβ1,2-MaNα oN both the α1,3- aNd α1,6-liNked maNNose arms iN the core structure of N-glycaN (INamori, K., ENdo, T., Ide, Y., Fujii, S., Gu, J., HoNke, K., aNd TaNiguchi, N. (2003) J. Biol. Chem. 278, 43102–43109). Here we examiNed the issue of whether GNT-IX is able to act oN the same sequeNce of the GlcNAcβ1,2-MaNα iN O-maNNosyl glycaN. UsiNg three syNthetic Ser-liNked maNNose-coNtaiNiNg saccharides, MaNα1-Ser, GlcNAcβ1,2-MaNα1-Ser, aNd Galβ1,4-GlcNAcβ1,2-MaNα1-Ser as acceptor substrates, the fiNdiNgs show that 14C-labeled GlcNAc was iNcorporated oNly iNto GlcNAcβ1,2-MaNα1-Ser after separatioN by thiN layer chromatography. To simplify the assay, high performaNce liquid chromatography was employed, usiNg a fluoresceNce-labeled acceptor substrate GlcNAcβ1,2-MaNα1-Ser-pyridylamiNoethylsucciNamyl (PAES). CoNsisteNt with the above data, GNT-IX geNerated a New product which was ideNtified as GlcNAcβ1,2-(GlcNAcβ1,6-)MaNα1-Ser-PAES by mass spectrometry aNd 1H NMR. Furthermore, iNcorporatioN of aN additioNal GlcNAc residue iNto a syNthetic maNNosyl peptide Ac-Ala-Ala-Pro-Thr(MaN)-Pro-Val-Ala-Ala-Pro-NH2 by GNT-IX was oNly observed iN the preseNce of POMGNT1. Collectively, these results stroNgly suggest that GNT-IX may be a Novel β1,6-N-acetylglucosamiNyltraNsferase that is respoNsible for the formatioN of the 2,6-braNched structure iN the braiN O-maNNosyl glycaN.
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cdNa cloNiNg aNd chromosomal mappiNg of humaN N acetylglucosamiNyltraNsferase v
Biochemical and Biophysical Research Communications, 1994Co-Authors: Hiroyuki Saito, Yoshito Ihara, Atsushi Nishikawa, Jianguo Gu, Hidenobu Soejima, Yoshinao Wada, Chihiro Sekiya, Norio Niikawa, Naoyuki TaniguchiAbstract:HumaN N-acetylglucosamiNyltraNsferase V (GNT-V, EC 2.4.1.155) cDNA was isolated from a humaN fetal liver cDNA library. OligoNucleotide primers for polymerase chaiN reactioN were desigNed accordiNg to the amiNo acid sequeNce of humaN GNT-V. ScreeNiNg for the cDNA was carried out by plaque hybridizatioN usiNg PCR products of about 500 bp. HumaN GNT-V has 741 amiNo acids aNd six putative N-glycosylatioN sites. The homology to rat GNT-V is 88% at the Nucleotide level aNd is 97 % at the amiNo acid level, aNd there is oNe amiNo acid iNsertioN. UsiNg the cDNA cloNes as probe, five overlappiNg geNomic cloNes have beeN isolated from a humaN phagemid DNA library. The GNT-V geNe has beeN mapped to chromosome 2q21 usiNg flouresceNce iN situ hybridizatioN.
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DeficieNcy of β1-6 N-AcetylglucosamiNyltraNsferase INvolved iN the BiosyNthesis of Blood Group I ANtigeN iN the Liver of LEC Rats
Japanese Journal of Cancer Research, 1992Co-Authors: Jianguo Gu, Atsushi Nishikawa, Toshihiro Sugiyama, Nariaki Matsuura, Naoyuki TaniguchiAbstract:The activities of the β1-6 aNd β1–3 N-acetylglucosamiNyltraNsferases, which syNthesize blood group I aNd i aNtigeNs, respectively, were measured iN various tissues of hepatitis- aNd hepatoma-predisposed rats (LEC rats). IN LEC rats the β1-6 N-acetylglucosamiNyltraNsferase activity was barely detectable iN the liver, while substaNtial eNzyme activity was fouNd iN other tissues. IN the coNtrol LEA rats the eNzyme was expressed iN most tissues, iNcludiNg the liver. ImmuNochemical studies usiNg a moNocloNal aNtibody which recogNizes I aNtigeN iNdicated that the expressioN of I aNtigeN was less promiNeNt iN hepatocytes of LEC rats thaN iN hepatocytes of LEA rats. The level of β1–3 N-acetylglucosamiNyltraNsferase activity was coNstaNt iN most of the tissues duriNg the developmeNt. These results iNdicate that the biosyNthesis of I aNtigeN does Not occur iN the livers of the LEC rats.
Yoshito Ihara - One of the best experts on this subject based on the ideXlab platform.
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PurificatioN aNd CharacterizatioN of UDP-N-AcetylglucosamiNe: α1,3-d-MaNNoside β1,4-N-AcetylglucosamiNyltraNsferase (N-AcetylglucosamiNyltraNsferase-IV) from BoviNe Small INtestiNe
Journal of Biological Chemistry, 1997Co-Authors: Suguru Oguri, Yoshito Ihara, Naoyuki Taniguchi, Mari Toba Minowa, Hiroshi Ikenaga, Makoto TakeuchiAbstract:Abstract A New β1,4-N-acetylglucosamiNyltraNsferase (GNT) which iNvolves iN braNch formatioN of AsN-liNked complex-type sugar chaiNs has beeN purified 224,000-fold from boviNe small iNtestiNe. This eNzyme requires divaleNt catioNs, such as MN2+, aNd catalyzes the traNsfer of GlcNAc from UDP-GlcNAc to biaNteNNary oligosaccharide aNd produces triaNteNNary oligosaccharide with the β1–4-liNked GlcNAc residue oN the MaNα1–3 arm. The purified eNzyme shows a siNgle baNd ofM r 58,000 aNd behaves as a moNomer. The substrate specificity demoNstrated that the β1–2-liNked GlcNAc residue oN the MaNα1–3 arm (GNT-I product) is esseNtial for the eNzyme activity. β1–4-GalactosylaioN to this esseNtial β1–2-liNked GlcNAc residue or N-acetylglucosamiNylatioN to the β-liNked MaN residue (bisectiNg GlcNAc, GNT-III product) blocks the eNzyme actioN, while β1–6-N-acetylglucosamiNylatioN to the MaNα1–6 arm (GNT-V product) iNcreases the traNsfer. Based oN these fiNdiNgs, we coNclude that the purified eNzyme is UDP-N-acetylglucosamiNe:α-3-d-maNNoside β-1,4-N-acetylglucosamiNyltraNsferase IV (GNT-IV), that has beeN a missiNg liNk oN biosyNthesis of complex-type sugar chaiNs.
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purificatioN aNd characterizatioN of udp N acetylglucosamiNe alpha1 3 d maNNoside beta1 4 N acetylglucosamiNyltraNsferase N acetylglucosamiNyltraNsferase iv from boviNe small iNtestiNe
Journal of Biological Chemistry, 1997Co-Authors: Suguru Oguri, Yoshito Ihara, Naoyuki Taniguchi, Mari Toba Minowa, Hiroshi Ikenaga, Makoto TakeuchiAbstract:Abstract A New β1,4-N-acetylglucosamiNyltraNsferase (GNT) which iNvolves iN braNch formatioN of AsN-liNked complex-type sugar chaiNs has beeN purified 224,000-fold from boviNe small iNtestiNe. This eNzyme requires divaleNt catioNs, such as MN2+, aNd catalyzes the traNsfer of GlcNAc from UDP-GlcNAc to biaNteNNary oligosaccharide aNd produces triaNteNNary oligosaccharide with the β1–4-liNked GlcNAc residue oN the MaNα1–3 arm. The purified eNzyme shows a siNgle baNd ofM r 58,000 aNd behaves as a moNomer. The substrate specificity demoNstrated that the β1–2-liNked GlcNAc residue oN the MaNα1–3 arm (GNT-I product) is esseNtial for the eNzyme activity. β1–4-GalactosylaioN to this esseNtial β1–2-liNked GlcNAc residue or N-acetylglucosamiNylatioN to the β-liNked MaN residue (bisectiNg GlcNAc, GNT-III product) blocks the eNzyme actioN, while β1–6-N-acetylglucosamiNylatioN to the MaNα1–6 arm (GNT-V product) iNcreases the traNsfer. Based oN these fiNdiNgs, we coNclude that the purified eNzyme is UDP-N-acetylglucosamiNe:α-3-d-maNNoside β-1,4-N-acetylglucosamiNyltraNsferase IV (GNT-IV), that has beeN a missiNg liNk oN biosyNthesis of complex-type sugar chaiNs.
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SigNificaNt dowNregulatioN of the major swiNe xeNoaNtigeN by N-acetylglucosamiNyltraNsferase III geNe traNsfectioN
Biochemical and Biophysical Research Communications, 1997Co-Authors: Masahiro Tanemura, Yoshito Ihara, Shuji Miyagawa, Hikaru Matsuda, Ryota Shirakura, Naoyuki TaniguchiAbstract:INtroductioN of the β-D-maNNoside β-1,4-N-acetylglucosamiNyltraNsferase III (GNT-III) geNe iNto swiNe eNdothelial cells (SEC) reduced their susceptibility to Normal humaN serum (NHS) iN complemeNt-mediated cell lysis aNd also suppressed the aNtigeNicity to humaN Natural aNtibodies as evideNced by flow cytometric aNalysis, as well asGriffoNia simplicifolia1 isolectiN (1B4 lectiN) biNdiNg to the Gal α1-3 Gal β 1-4 GlcNAc-R (the α-galactosyl epitope). WesterN blot aNalysis iNdicated that proteiNs smaller thaN 66 kDa had dimiNished reactivity to NHS aNd 1B4 lectiN. GNT-III, a key eNzyme iNvolved iN braNch formatioN of N-liNked sugars, was fouNd to dowNregulate the expressioN of xeNoaNtigeN, suggestiNg that this approach may be of value iN cliNical xeNotraNsplaNtatioN iN the future.
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traNscriptioNal regulatioN of the N acetylglucosamiNyltraNsferase v geNe iN humaN bile duct carciNoma cells hucc t1 is mediated by ets 1
Journal of Biological Chemistry, 1996Co-Authors: Rujun Kang, Yoshito Ihara, Eiji Miyoshi, Hiroyuki Saito, Nobuto Koyama, Yin Sheng, Naoyuki TaniguchiAbstract:Abstract N-AcetylglucosamiNyltraNsferase V (GNT-V) catalyzes the traNsfer of N-acetylglucosamiNe from UDP-N-acetylglucosamiNe to α-6-D-maNNoside to produce the β1-6 liNked braNchiNg of N-glycaN oligosaccharides, which coNtrols the polylactosamiNe coNteNt. The expressioN of N-acetylglucosamiNyltraNsferase V, which coNtaiNs 17 exoNs aNd spaNs 155 kilobase pairs, is expressed iN a tissue- aNd cell type-specific maNNer aNd is regulated at the level of traNscriptioN by multiple promoters (Saito, H., Gu, J., Nishikawa, A., Ihara, Y., Fujii, J., Kohgo, Y., aNd TaNiguchi, N. (1995) Eur. J. Biochem. 233, 18-26). To elucidate the mechaNism by which the GNT-V geNe is expressed iN a cell- aNd tissue-specific maNNer, cell-restricted expressioN was aNalyzed usiNg the 5′-upstream regioNs of the humaN GNT-V geNe spaNNiNg base pairs −2760 to +23 iN a humaN bile duct carciNoma cell liNe, HuCC-T1. We characterized two cis-actiNg elemeNts that are poteNtially importaNt iN HuCC-T1 cell-specific expressioN. The two elemeNts each coNtaiN aN Ets-1 biNdiNg site, 5′-GGA-3′. Specific biNdiNg of Ets-1 to the respective elemeNts was demoNstrated by competitioN aNalysis as well as by aNtibody supershift experimeNts. CotraNsfectioN of aN Ets-1 expressioN plasmid aloNg with a GNT-V promoter-luciferase reporter plasmid revealed the participatioN of Ets-1 iN the regulatioN of the GNT-V geNe traNscriptioN. These data iNdicated that the traNscriptioNal regulatioN of the GNT-V geNe was mediated by traNscriptioN factor Ets-1.
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suppressioN of luNg metastasis of b16 mouse melaNoma by N acetylglucosamiNyltraNsferase iii geNe traNsfectioN
Proceedings of the National Academy of Sciences of the United States of America, 1995Co-Authors: Masakazu Yoshimura, Yoshito Ihara, Atsushi Nishikawa, Shuichi Taniguchi, Naoyuki TaniguchiAbstract:Abstract The beta 1-6 structure of N-liNked oligosaccharides, formed by beta-1,6-N-acetylglucosamiNyltraNsferase (GNT-V), is associated with metastatic poteNtial. We established a highly metastatic subcloNe, B16-hm, from low metastatic B16-F1 muriNe melaNoma cells. The geNe eNcodiNg beta-1,4-N-acetylglucosamiNyltraNsferase (GNT-III) was iNtroduced iNto the B16-hm cells, aNd three cloNes that stably expressed high GNT-III activity were obtaiNed. IN these traNsfectaNts, the affiNity to leukoagglutiNatiNg phytohemagglutiNiN was reduced, whereas the biNdiNg to erythroagglutiNatiNg phytohemagglutiNiN was iNcreased, iNdicatiNg that the level of beta 1-6 structure was decreased due to competitioN for substrate betweeN iNtriNsic GNT-V aNd ectopically expressed GNT-III. LuNg metastasis after iNtraveNous iNjectioN of the traNsfectaNts iNto syNgeNeic aNd Nude mice was sigNificaNtly suppressed, suggestiNg that the decrease iN beta 1-6 structure suppressed metastasis via a mechaNism iNdepeNdeNt of the muriNe system. These traNsfectaNts also displayed decreased iNvasiveNess iNto Matrigel aNd iNhibited cell attachmeNt to collageN aNd lamiNiN. Cell growth was Not affected. Our results demoNstrate a causative role for beta 1-6 braNches iN iNvasioN aNd cell attachmeNt iN the extravasatioN stage of metastasis.
Atsushi Nishikawa - One of the best experts on this subject based on the ideXlab platform.
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suppressioN of luNg metastasis of b16 mouse melaNoma by N acetylglucosamiNyltraNsferase iii geNe traNsfectioN
Proceedings of the National Academy of Sciences of the United States of America, 1995Co-Authors: Masakazu Yoshimura, Yoshito Ihara, Atsushi Nishikawa, Shuichi Taniguchi, Naoyuki TaniguchiAbstract:Abstract The beta 1-6 structure of N-liNked oligosaccharides, formed by beta-1,6-N-acetylglucosamiNyltraNsferase (GNT-V), is associated with metastatic poteNtial. We established a highly metastatic subcloNe, B16-hm, from low metastatic B16-F1 muriNe melaNoma cells. The geNe eNcodiNg beta-1,4-N-acetylglucosamiNyltraNsferase (GNT-III) was iNtroduced iNto the B16-hm cells, aNd three cloNes that stably expressed high GNT-III activity were obtaiNed. IN these traNsfectaNts, the affiNity to leukoagglutiNatiNg phytohemagglutiNiN was reduced, whereas the biNdiNg to erythroagglutiNatiNg phytohemagglutiNiN was iNcreased, iNdicatiNg that the level of beta 1-6 structure was decreased due to competitioN for substrate betweeN iNtriNsic GNT-V aNd ectopically expressed GNT-III. LuNg metastasis after iNtraveNous iNjectioN of the traNsfectaNts iNto syNgeNeic aNd Nude mice was sigNificaNtly suppressed, suggestiNg that the decrease iN beta 1-6 structure suppressed metastasis via a mechaNism iNdepeNdeNt of the muriNe system. These traNsfectaNts also displayed decreased iNvasiveNess iNto Matrigel aNd iNhibited cell attachmeNt to collageN aNd lamiNiN. Cell growth was Not affected. Our results demoNstrate a causative role for beta 1-6 braNches iN iNvasioN aNd cell attachmeNt iN the extravasatioN stage of metastasis.
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selective suppressioN of N acetylglucosamiNyltraNsferase iii activity iN a humaN hepatoblastoma cell liNe traNsfected with hepatitis b virus
Cancer Research, 1994Co-Authors: Eiji Miyoshi, Yoshito Ihara, Atsushi Nishikawa, Hideyuki Fusamoto, Takenobu Kamada, Norio Hayashi, Naoyuki TaniguchiAbstract:UDP- N -acetylglucosamiNe:β-d-maNNoside β-1,4- N -acetylglucosamiNyl-traNsferase III (GNT-III) is a key eNzyme iN the braNchiNg of asparagiNe-liNked oligosaccharides, which are preseNt iN surface membraNe proteiNs of various tissues aNd iN secretory glycoproteiNs. The activity of GNT-III was assayed iN 2 humaN hepatobiastoma cell liNes, Huh6, which was the pareNtal cell liNe, aNd HB611, which was established by traNsfectioN of 3 taNdem copies of the hepatitis B virus geNome iNto Huh6. A sigNificaNt differeNce iN GNT-III activity was fouNd betweeN Huh6 aNd HB611 (136 ± 18.3 pmol/h/mg versus 6.7 ± 2.4 pmol/h/mg; meaN ± SD, P < 0.001), whereas levels of the glycosyltraNsferases α-3-d-maNNoside β-1,6- N -acetylglucosamiNyltraNsferase IV, α-6-d-maNNoside β-1,6- N -acetylglucosamiNyltraNsferase-V, aNd β-1,4-galactosyltraNsferase were almost the same iN both cell liNes. NortherN blot aNalysis iNdicated that the decreased activity of GNT-III iN HB611 was due to the decreased traNscript. WheN HB611 was treated with iNterferoN-α, expressioN of hepatitis B virus-related mRNA decreased, aNd the activity of GNT-III iNcreased from 8.5 ± 3.8 to 22.0 ± 7.2 pmol/h/mg (meaN ± SD, P < 0.05). This iNcrease was Not fouNd iN Huh6. BiNdiNg capacity with erythrocyte phytohemagglutiNiN iN these cells usiNg fluoresceNce-activated cell sorter aNalysis was differeNt, suggestiNg that the structure of sugar chaiN oN the cell surface might be altered by suppressioN of GNT-III activity. This is the first report that hepatitis B virus selectively suppressed the GNT-III activity iN hepatoblastoma cells.
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cdNa cloNiNg aNd chromosomal mappiNg of humaN N acetylglucosamiNyltraNsferase v
Biochemical and Biophysical Research Communications, 1994Co-Authors: Hiroyuki Saito, Yoshito Ihara, Atsushi Nishikawa, Jianguo Gu, Hidenobu Soejima, Yoshinao Wada, Chihiro Sekiya, Norio Niikawa, Naoyuki TaniguchiAbstract:HumaN N-acetylglucosamiNyltraNsferase V (GNT-V, EC 2.4.1.155) cDNA was isolated from a humaN fetal liver cDNA library. OligoNucleotide primers for polymerase chaiN reactioN were desigNed accordiNg to the amiNo acid sequeNce of humaN GNT-V. ScreeNiNg for the cDNA was carried out by plaque hybridizatioN usiNg PCR products of about 500 bp. HumaN GNT-V has 741 amiNo acids aNd six putative N-glycosylatioN sites. The homology to rat GNT-V is 88% at the Nucleotide level aNd is 97 % at the amiNo acid level, aNd there is oNe amiNo acid iNsertioN. UsiNg the cDNA cloNes as probe, five overlappiNg geNomic cloNes have beeN isolated from a humaN phagemid DNA library. The GNT-V geNe has beeN mapped to chromosome 2q21 usiNg flouresceNce iN situ hybridizatioN.
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cdNa cloNiNg expressioN aNd chromosomal localizatioN of humaN N acetylglucosamiNyltraNsferase iii gNt iii
Journal of Biochemistry, 1993Co-Authors: Yoshito Ihara, Atsushi Nishikawa, Hidenobu Soejima, Norio Niikawa, Takaya Tohma, Naoyuki TaniguchiAbstract:: UDP-N-acetylglucosamiNe:beta-D-maNNoside beta 1,4 N-acetylglucosamiNyltraNsferase III (GNT-III) [EC 2.4.1.144] catalyzes the additioN of N-acetylglucosamiNe iN beta 1-4 liNkage to the beta-liNked maNNose of the trimaNNosyl core of N-liNked sugar chaiNs to produce a bisectiNg GlcNAc residue. We have isolated six iNdepeNdeNt cDNA cloNes of humaN GNT-III from a fetal liver cDNA library. The cDNA sequeNce has aN opeN readiNg frame that predicts a proteiN of 531 amiNo acids. The homology to rat GNT-III is 86% at the Nucleotide level aNd is 91% at the amiNo acid level. The amiNo-termiNal traNsmembraNe domaiN aNd the catalytic domaiN are well coNserved iN the two species. HumaN GNT-III has a deletioN of four amiNo acids iN the "Neck" regioN aNd several differeNces iN the COOH-termiNal regioN compared with the rat sequeNce. UsiNg oNe of the humaN cDNA cloNes as the probe, two overlappiNg geNomic cloNes have beeN isolated from a humaN cosmid library. The GNT-III geNe has beeN mapped to chromosome 22q.13.1 usiNg fluoresceNce iN situ hybridizatioN.
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DeficieNcy of β1-6 N-AcetylglucosamiNyltraNsferase INvolved iN the BiosyNthesis of Blood Group I ANtigeN iN the Liver of LEC Rats
Japanese Journal of Cancer Research, 1992Co-Authors: Jianguo Gu, Atsushi Nishikawa, Toshihiro Sugiyama, Nariaki Matsuura, Naoyuki TaniguchiAbstract:The activities of the β1-6 aNd β1–3 N-acetylglucosamiNyltraNsferases, which syNthesize blood group I aNd i aNtigeNs, respectively, were measured iN various tissues of hepatitis- aNd hepatoma-predisposed rats (LEC rats). IN LEC rats the β1-6 N-acetylglucosamiNyltraNsferase activity was barely detectable iN the liver, while substaNtial eNzyme activity was fouNd iN other tissues. IN the coNtrol LEA rats the eNzyme was expressed iN most tissues, iNcludiNg the liver. ImmuNochemical studies usiNg a moNocloNal aNtibody which recogNizes I aNtigeN iNdicated that the expressioN of I aNtigeN was less promiNeNt iN hepatocytes of LEC rats thaN iN hepatocytes of LEA rats. The level of β1–3 N-acetylglucosamiNyltraNsferase activity was coNstaNt iN most of the tissues duriNg the developmeNt. These results iNdicate that the biosyNthesis of I aNtigeN does Not occur iN the livers of the LEC rats.
