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<title>Enzyme assay of N-acetylglucosamine-6-O-sulfotransferases - Glycoscience Protocols (GlycoPODv2) - NCBI Bookshelf</title>
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<meta name="robots" content="INDEX,FOLLOW,NOARCHIVE" /><meta name="citation_inbook_title" content="Glycoscience Protocols (GlycoPODv2) [Internet]" /><meta name="citation_title" content="Enzyme assay of N-acetylglucosamine-6-O-sulfotransferases" /><meta name="citation_publisher" content="Japan Consortium for Glycobiology and Glycotechnology" /><meta name="citation_date" content="2022/03/18" /><meta name="citation_author" content="Kenji Uchimura" /><meta name="citation_pmid" content="37590713" /><meta name="citation_fulltext_html_url" content="https://www.ncbi.nlm.nih.gov/books/NBK593984/" /><meta name="citation_keywords" content="carbohydrate sulfotransferase" /><meta name="citation_keywords" content="N-acetylglucosamine-6-O-sulfotransferase" /><meta name="citation_keywords" content="enzyme assay" /><meta name="citation_keywords" content="thin-layer chromatography" /><meta name="citation_keywords" content="selectin ligand" /><meta name="citation_keywords" content="keratan sulfate" /><meta name="citation_keywords" content="sulfated N-acetyllactosamine" /><link rel="schema.DC" href="http://purl.org/DC/elements/1.0/" /><meta name="DC.Title" content="Enzyme assay of N-acetylglucosamine-6-O-sulfotransferases" /><meta name="DC.Type" content="Text" /><meta name="DC.Publisher" content="Japan Consortium for Glycobiology and Glycotechnology" /><meta name="DC.Contributor" content="Kenji Uchimura" /><meta name="DC.Date" content="2022/03/18" /><meta name="DC.Identifier" content="https://www.ncbi.nlm.nih.gov/books/NBK593984/" /><meta name="description" content="N-Acetylglucosamine-6-O-sulfotransferase (GlcNAc6ST) transfers a sulfate group from 3’-phosphoadenosine 5’-phosphosulfate (PAPS) to C-6 of N-acetylglucosamine (GlcNAc) exposed at the nonreducing end of glycans (1). The discovery of the GlcNAc6ST gene was reported in 1998 (1). Five members of the GlcNAc6ST family have been identified in humans, four of which have mouse orthologs so far (2). GlcNAc6ST1 (encoded by Chst2) (1, 3) and GlcNAc6ST2 (Chst4) (4–6) are involved in the synthesis of sialyl 6-sulfo Lewis X, a major class of L-selectin ligands (7, 8). GlcNAc6ST3 (Chst5), originally found as an intestinal GlcNAc6ST (9), is also known as a keratan sulfate (KS) GlcNAc6ST in adult brains (10). GlcNAc6ST1 mediates synthesis of KS in developing brains (11, 12) and microglial KS-related N-acetyllactosamine (LacNAc) oligosaccharides in neurodegenerative diseases (13–15). GlcNAc6ST4 (Chst7) has the highest homology with GlcNAc6ST1 compared to other members and is capable of sulfating GalNAc with a weak activity (16–18). GlcNAc6ST5 (CHST6) is a human corneal enzyme synthesizing KS. Defective CHST6 causes macular corneal dystrophy (19). We describe here an in vitro assay for GlcNAc6STs. Recombinant GlcNAc6STs and various oligosaccharides derived from N-linked or O-linked glycans can be used in this assay." /><meta name="og:title" content="Enzyme assay of N-acetylglucosamine-6-O-sulfotransferases" /><meta name="og:type" content="book" /><meta name="og:description" content="N-Acetylglucosamine-6-O-sulfotransferase (GlcNAc6ST) transfers a sulfate group from 3’-phosphoadenosine 5’-phosphosulfate (PAPS) to C-6 of N-acetylglucosamine (GlcNAc) exposed at the nonreducing end of glycans (1). The discovery of the GlcNAc6ST gene was reported in 1998 (1). Five members of the GlcNAc6ST family have been identified in humans, four of which have mouse orthologs so far (2). GlcNAc6ST1 (encoded by Chst2) (1, 3) and GlcNAc6ST2 (Chst4) (4–6) are involved in the synthesis of sialyl 6-sulfo Lewis X, a major class of L-selectin ligands (7, 8). GlcNAc6ST3 (Chst5), originally found as an intestinal GlcNAc6ST (9), is also known as a keratan sulfate (KS) GlcNAc6ST in adult brains (10). GlcNAc6ST1 mediates synthesis of KS in developing brains (11, 12) and microglial KS-related N-acetyllactosamine (LacNAc) oligosaccharides in neurodegenerative diseases (13–15). GlcNAc6ST4 (Chst7) has the highest homology with GlcNAc6ST1 compared to other members and is capable of sulfating GalNAc with a weak activity (16–18). GlcNAc6ST5 (CHST6) is a human corneal enzyme synthesizing KS. Defective CHST6 causes macular corneal dystrophy (19). We describe here an in vitro assay for GlcNAc6STs. Recombinant GlcNAc6STs and various oligosaccharides derived from N-linked or O-linked glycans can be used in this assay." /><meta name="og:url" content="https://www.ncbi.nlm.nih.gov/books/NBK593984/" /><meta name="og:site_name" content="NCBI Bookshelf" /><meta name="og:image" content="https://www.ncbi.nlm.nih.gov/corehtml/pmc/pmcgifs/bookshelf/thumbs/th-glycopodv2-lrg.png" /><meta name="twitter:card" content="summary" /><meta name="twitter:site" content="@ncbibooks" /><meta name="bk-non-canon-loc" content="/books/n/glycopodv2/g83-assay6st/" /><link rel="canonical" href="https://www.ncbi.nlm.nih.gov/books/NBK593984/" /><link rel="stylesheet" href="/corehtml/pmc/css/figpopup.css" type="text/css" media="screen" /><link rel="stylesheet" href="/corehtml/pmc/css/bookshelf/2.26/css/books.min.css" type="text/css" /><link rel="stylesheet" href="/corehtml/pmc/css/bookshelf/2.26/css/books_print.min.css" type="text/css" /><style type="text/css">p a.figpopup{display:inline !important} .bk_tt {font-family: monospace} .first-line-outdent .bk_ref {display: inline} </style><script type="text/javascript" src="/corehtml/pmc/js/jquery.hoverIntent.min.js"> </script><script type="text/javascript" src="/corehtml/pmc/js/common.min.js?_=3.18"> </script><script type="text/javascript">window.name="mainwindow";</script><script type="text/javascript" src="/corehtml/pmc/js/bookshelf/2.26/book-toc.min.js"> </script><script type="text/javascript" src="/corehtml/pmc/js/bookshelf/2.26/books.min.js"> </script>
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<div class="pre-content"><div><div class="bk_prnt"><p class="small">NCBI Bookshelf. A service of the National Library of Medicine, National Institutes of Health.</p><p>Nishihara S, Angata K, Aoki-Kinoshita KF, et al., editors. Glycoscience Protocols (GlycoPODv2) [Internet]. Saitama (JP): Japan Consortium for Glycobiology and Glycotechnology; 2021-. </p></div></div></div>
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<div class="main-content lit-style" itemscope="itemscope" itemtype="http://schema.org/CreativeWork"><div class="meta-content fm-sec"><h1 id="_NBK593984_"><span class="title" itemprop="name">Enzyme assay of <i>N</i>-acetylglucosamine-6-<i>O</i>-sulfotransferases</span></h1><div class="contrib half_rhythm"><span itemprop="author">Kenji Uchimura</span>, Ph.D.<div class="affiliation small">Unité de Glycobiologie Structurale et Fonctionnelle, CNRS, UMR8576, Université de Lille, France<div><span class="email-label">Email: </span><a href="mailto:dev@null" data-email="rf.ellil-vinu@arumihcu.ijnek" class="oemail">rf.ellil-vinu@arumihcu.ijnek</a></div></div><div class="small">Corresponding author.</div></div><p class="small">Created: <span itemprop="datePublished">September 26, 2021</span>; Last Revision: <span itemprop="dateModified">March 18, 2022</span>.</p></div><div class="body-content whole_rhythm" itemprop="text"><div id="g83-assay6st.Introduction"><h2 id="_g83-assay6st_Introduction_">Introduction:</h2><p><i>N</i>-Acetylglucosamine-6-<i>O</i>-sulfotransferase (GlcNAc6ST) transfers a sulfate group from 3’-phosphoadenosine 5’-phosphosulfate (PAPS) to C-6 of <i>N</i>-acetylglucosamine (GlcNAc) exposed at the nonreducing end of glycans (<a class="bk_pop" href="#g83-assay6st.REF.1">1</a>). The discovery of the GlcNAc6ST gene was reported in 1998 (<a class="bk_pop" href="#g83-assay6st.REF.1">1</a>). Five members of the GlcNAc6ST family have been identified in humans, four of which have mouse orthologs so far (<a class="bk_pop" href="#g83-assay6st.REF.2">2</a>). GlcNAc6ST1 (encoded by <i>Chst2</i>) (<a class="bk_pop" href="#g83-assay6st.REF.1">1</a>, <a class="bk_pop" href="#g83-assay6st.REF.3">3</a>) and GlcNAc6ST2 (<i>Chst4</i>) (<a class="bk_pop" href="#g83-assay6st.REF.4">4</a>–<a class="bk_pop" href="#g83-assay6st.REF.6">6</a>) are involved in the synthesis of sialyl 6-sulfo Lewis X, a major class of L-selectin ligands (<a class="bk_pop" href="#g83-assay6st.REF.7">7</a>, <a class="bk_pop" href="#g83-assay6st.REF.8">8</a>). GlcNAc6ST3 (<i>Chst5</i>), originally found as an intestinal GlcNAc6ST (<a class="bk_pop" href="#g83-assay6st.REF.9">9</a>), is also known as a keratan sulfate (KS) GlcNAc6ST in adult brains (10). GlcNAc6ST1 mediates synthesis of KS in developing brains (<a class="bk_pop" href="#g83-assay6st.REF.11">11</a>, <a class="bk_pop" href="#g83-assay6st.REF.12">12</a>) and microglial KS-related <i>N</i>-acetyllactosamine (LacNAc) oligosaccharides in neurodegenerative diseases (<a class="bk_pop" href="#g83-assay6st.REF.13">13</a>–<a class="bk_pop" href="#g83-assay6st.REF.15">15</a>). GlcNAc6ST4 (<i>Chst7</i>) has the highest homology with GlcNAc6ST1 compared to other members and is capable of sulfating GalNAc with a weak activity (<a class="bk_pop" href="#g83-assay6st.REF.16">16</a>–<a class="bk_pop" href="#g83-assay6st.REF.18">18</a>). GlcNAc6ST5 (<i>CHST6</i>) is a human corneal enzyme synthesizing KS. Defective <i>CHST6</i> causes macular corneal dystrophy (<a class="bk_pop" href="#g83-assay6st.REF.19">19</a>). We describe here an <i>in vitro</i> assay for GlcNAc6STs. Recombinant GlcNAc6STs and various oligosaccharides derived from <i>N</i>-linked or <i>O</i>-linked glycans can be used in this assay.</p></div><div id="g83-assay6st.Protocol"><h2 id="_g83-assay6st_Protocol_">Protocol</h2><p>The following protocol of enzyme assay is based on thin-layer chromatography (TLC) system. TLC is suitable for handling numerous samples. Oligosaccharide substrates tested are GlcNAcβ1-6Man-<i>O</i>-methyl, GlcNAcβ1-2Man, GlcNAcβ1-6[Galβ1-3]GalNAc-<i>p</i>-nitrophenyl (Core2-pNP), GlcNAcβ1-3GalNAc-<i>p</i>-nitrophenyl (Core3-pNP), and GlcNAcβ1-3Galβ1-4GlcNAcβ1-3Galβ1-4GlcNAc (<a class="bk_pop" href="#g83-assay6st.REF.17">17</a>, <a class="bk_pop" href="#g83-assay6st.REF.20">20</a>) (<a class="figpopup" href="/books/NBK593984/table/g83-assay6st.T.comparison_of_the_substra/?report=objectonly" target="object" rid-figpopup="figg83assay6stTcomparisonofthesubstra" rid-ob="figobg83assay6stTcomparisonofthesubstra">Table 1</a>).</p><div id="g83-assay6st.Materials"><h3>Materials</h3><dl class="temp-labeled-list"><dt>1.</dt><dd><p class="no_top_margin">Phosphate-buffered saline (PBS)</p></dd><dt>2.</dt><dd><p class="no_top_margin">Ham’s F12 modified medium (Thermo Fisher Scientific, Waltham, MA, USA)</p></dd><dt>3.</dt><dd><p class="no_top_margin">Dulbecco’s modified Eagle’s medium (DMEM; Thermo Fisher Scientific)</p></dd><dt>4.</dt><dd><p class="no_top_margin">Fetal bovine serum (FBS; Thermo Fisher Scientific)</p></dd><dt>5.</dt><dd><p class="no_top_margin">Chinese Hamster ovary (CHO) cells (CHO-K1; ATCC, Manassas, VA, USA)</p></dd><dt>6.</dt><dd><p class="no_top_margin">COS-7 cells (simian kidney cell line; ATCC)</p></dd><dt>7.</dt><dd><p class="no_top_margin">Lipofectamine™ 2000 (Thermo Fisher Scientific)</p></dd><dt>8.</dt><dd><p class="no_top_margin">1M Tris-HCl, pH 7.5</p></dd><dt>9.</dt><dd><p class="no_top_margin">3M Sucrose</p></dd><dt>10.</dt><dd><p class="no_top_margin">Immunoglobulin G (IgG)-Sepharose 6 Fast Flow (Cytiva, Marlborough, MA, USA)</p></dd><dt>11.</dt><dd><p class="no_top_margin">10% (wt/vol) Triton X-100 solution</p></dd><dt>12.</dt><dd><p class="no_top_margin">80% (vol/vol) glycerol solution</p></dd><dt>13.</dt><dd><p class="no_top_margin">50 mM of adenosine 5’-monophosphate (AMP)</p></dd><dt>14.</dt><dd><p class="no_top_margin">1M sodium fluoride (NaF)</p></dd><dt>15.</dt><dd><p class="no_top_margin">0.2M MnCl2</p></dd><dt>16.</dt><dd><p class="no_top_margin">Thick-wall ultracentrifuge open tubes (Beckman Coulter, Brea, CA, USA)</p></dd><dt>17.</dt><dd><p class="no_top_margin">TLC aluminum plates coated with cellulose (0.1-mm thick; Merck, Darmstadt, Germany)</p></dd><dt>18.</dt><dd><p class="no_top_margin">TLC-developing solvent: ethanol/pyridine/n-butyl alcohol/water/acetate (100:10:10:30:3 by volume)</p></dd><dt>19.</dt><dd><p class="no_top_margin">[<sup>35</sup>S]-PAPS (1.9 Ci/mmol; PerkinElmer, Waltham, MA, USA)</p></dd></dl></div><div id="g83-assay6st.Instruments"><h3>Instruments</h3><dl class="temp-labeled-list"><dt>1.</dt><dd><p class="no_top_margin">Tube rotator (Thermo Fisher Scientific)</p></dd><dt>2.</dt><dd><p class="no_top_margin">Optima TLX Ultracentrifuge (Beckman Coulter)</p></dd><dt>3.</dt><dd><p class="no_top_margin">TLC developing tank (Merck)</p></dd><dt>4.</dt><dd><p class="no_top_margin">BAS2000 bioimaging analyzer (Fuji Film, Tokyo, Japan) (<b>Note 1</b>)</p></dd></dl></div><div id="g83-assay6st.Methods"><h3>Methods</h3><dl class="temp-labeled-list"><dt>1.</dt><dd><p class="no_top_margin">Expression of GlcNAc6ST in cultured cells and collect the enzyme as a microsome fraction</p><dl class="temp-labeled-list"><dt>a.</dt><dd><p class="no_top_margin">Construction of expression plasmids for full-length GlcNAc6STs is described previously (<a class="bk_pop" href="#g83-assay6st.REF.1">1</a>, <a class="bk_pop" href="#g83-assay6st.REF.4">4</a>, <a class="bk_pop" href="#g83-assay6st.REF.6">6</a>, <a class="bk_pop" href="#g83-assay6st.REF.9">9</a>, <a class="bk_pop" href="#g83-assay6st.REF.17">17</a>, <a class="bk_pop" href="#g83-assay6st.REF.21">21</a>).</p></dd><dt>b.</dt><dd><p class="no_top_margin">Culture CHO cells on 10-cm dishes and passage them until they reach ~70% confluence with Ham’s F12 modified medium containing 10% FBS.</p></dd><dt>c.</dt><dd><p class="no_top_margin">Rinse the cells with warm PBS and then transfect transiently with a GlcNAc6ST expression plasmid using Lipofectamine™ 2000 according to the manufacturer’s instructions.</p></dd><dt>d.</dt><dd><p class="no_top_margin">Culture the cells in Ham’s F12 modified medium containing 10% FBS for 48 h.</p></dd><dt>e.</dt><dd><p class="no_top_margin">Wash the cell layer with PBS, scrape the cells off the dish, and homogenize them in 3 mL of 0.25 M sucrose and 20 mM of Tris-HCl, pH 7.5.</p></dd><dt>f.</dt><dd><p class="no_top_margin">Centrifuge the homogenates briefly and then transfer the supernatant to an ultracentrifuge tube.</p></dd><dt>g.</dt><dd><p class="no_top_margin">Ultracentrifuge the tube at 100,000 ×<i>g</i> for 1 h.</p></dd><dt>h.</dt><dd><p class="no_top_margin">Discard the supernatant and then dissolve the precipitate in 50 µL of 20 mM of Tris-HCl, pH 7.5, 0.5% Triton X-100, and 10% glycerol.</p></dd><dt>i.</dt><dd><p class="no_top_margin">Use this microsome fraction as the enzyme source.</p></dd></dl></dd><dt>2.</dt><dd><p class="no_top_margin">Expression and purification of GlcNAc6STs fused with protein A</p><dl class="temp-labeled-list"><dt>a.</dt><dd><p class="no_top_margin">Construction of the plasmids encoding fusion proteins of GlcNAc6STs to the IgM signal sequence and protein A in the N-terminal region are as described (<a class="bk_pop" href="#g83-assay6st.REF.20">20</a>).</p></dd><dt>b.</dt><dd><p class="no_top_margin">Culture COS-7 cells on 10-cm dishes and passage them until they reach ~70% confluence with DMEM containing 10% FBS.</p></dd><dt>c.</dt><dd><p class="no_top_margin">Rinse the cells with warm PBS and then transfect transiently with protein A-fused GlcNAc6ST expression plasmid using Lipofectamine™ 2000 according to the manufacturer’s instructions.</p></dd><dt>d.</dt><dd><p class="no_top_margin">Replace the medium with DMEM containing 2% IgG-free FBS (or IgG ultra-low level FBS), and then culture the cells for 48 h (<b>Note 2</b>).</p></dd><dt>e.</dt><dd><p class="no_top_margin">Collect 10 mL of conditioned medium (CM) and mix with 10 µL of IgG-Sepharose resin in a tube.</p></dd><dt>f.</dt><dd><p class="no_top_margin">Incubate the tube at 4°C for 3 h with gently rocking to adsorb the protein A fused-GlcNAc6STs in CM to the resin.</p></dd><dt>g.</dt><dd><p class="no_top_margin">Collect the resin by centrifugation at 12,000 ×<i>g</i> and then wash three times with PBS.</p></dd><dt>h.</dt><dd><p class="no_top_margin">Suspend the resin in 50 µL of 50 mM of Tris-HCl, pH 7.5, and used as enzymes.</p></dd></dl></dd><dt>3.</dt><dd><p class="no_top_margin">Assay of GlcNAc6ST activities toward oligosaccharides</p><dl class="temp-labeled-list"><dt>a.</dt><dd><p class="no_top_margin">Prepare the standard reaction mix in a 1.5-mL tube containing 1 µmol of Tris-HCl, pH7.5, 0.2 µmol of MnCl<sub>2</sub>, 0.04 µmol of AMP, 2 µmol of NaF, 20 nmol of oligosaccharides, 150 pmol of [<sup>35</sup>S]-PAPS (1.5 × 10<sup>6</sup> cpm), 0.05% Triton X-100, and 1 µL of microsome fraction or fusion protein suspension in a final volume of 20 µL.</p></dd><dt>b.</dt><dd><p class="no_top_margin">Incubate the tube containing the reaction mix at 30°C for 1 h.</p></dd><dt>c.</dt><dd><p class="no_top_margin">Apply aliquots of 2 µL of the reaction mix to TLC plates.</p></dd><dt>d.</dt><dd><p class="no_top_margin">Develop the plates with the developing solvent.</p></dd><dt>e.</dt><dd><p class="no_top_margin">End the development when the solvent front reaches the top of the plates. The <sup>35</sup>S-labeled oligosaccharides migrate faster than [<sup>35</sup>S]-PAPS.</p></dd><dt>f.</dt><dd><p class="no_top_margin">Visualize and measure the radioactivity of the <sup>35</sup>S-labeled oligosaccharides with a BAS2000 bioimaging analyzer (<b>Note 3</b>).</p></dd></dl></dd></dl></div><div id="g83-assay6st.Notes"><h3>Notes</h3><dl class="temp-labeled-list"><dt>1.</dt><dd><p class="no_top_margin">Other devices that allow for conducting auto-radiography with imaging plates and quantify the radioactivity of the image recorded on a plate are also useful instead of the BAS system.</p></dd><dt>2.</dt><dd><p class="no_top_margin">IgG-free or IgG ultra-low level FBS can be prepared by applying FBS to protein-G affinity column chromatography. IgG ultra-low level FBS is also commercially available (Thermo Fisher Scientific).</p></dd><dt>3.</dt><dd><p class="no_top_margin">The reaction product can also be analyzed using Superdex 30 chromatography (<a class="bk_pop" href="#g83-assay6st.REF.1">1</a>, <a class="bk_pop" href="#g83-assay6st.REF.17">17</a>).</p></dd></dl></div></div><div id="g83-assay6st.References"><h2 id="_g83-assay6st_References_">References</h2><dl class="temp-labeled-list"><dt>1.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.1">Uchimura K, Muramatsu H, Kadomatsu K, Fan QW, Kurosawa N, Mitsuoka C, Kannagi R, Habuchi O, Muramatsu T. Molecular cloning and characterization of an <em>N</em>-acetylglucosamine-6-<em>O</em>-sulfotransferase. <span><span class="ref-journal">J Biol Chem. </span>1998;<span class="ref-vol">273</span>(35):22577–83.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/9712885" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 9712885</span></a>] [<a href="http://dx.crossref.org/10.1074/jbc.273.35.22577" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>2.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.2">Uchimura K, Rosen SD. Sulfated L-selectin ligands as a therapeutic target in chronic inflammation. <span><span class="ref-journal">Trends Immunol. </span>2006;<span class="ref-vol">27</span>(12):559–65.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/17049924" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 17049924</span></a>] [<a href="http://dx.crossref.org/10.1016/j.it.2006.10.007" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>3.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.3">Uchimura K, Kadomatsu K, El-Fasakhany FM, Singer MS, Izawa M, Kannagi R, Takeda N, Rosen SD, Muramatsu T. N-acetylglucosamine 6-<em>O</em>-sulfotransferase-1 regulates expression of L-selectin ligands and lymphocyte homing. <span><span class="ref-journal">J Biol Chem. </span>2004;<span class="ref-vol">279</span>(33):35001–8.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/15175329" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 15175329</span></a>] [<a href="http://dx.crossref.org/10.1074/jbc.M404456200" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>4.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.4">Bistrup A, Bhakta S, Lee JK, Belov YY, Gunn MD, Zuo FR, Huang CC, Kannagi R, Rosen SD. Sulfotransferases of two specificities function in the reconstitution of high endothelial cell ligands for L-selectin. <span><span class="ref-journal">J Cell Biol. </span>1999;<span class="ref-vol">145</span>(4):899–910.</span> [<a href="/pmc/articles/PMC2133194/" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pmc">PMC free article<span class="bk_prnt">: PMC2133194</span></a>] [<a href="https://pubmed.ncbi.nlm.nih.gov/10330415" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 10330415</span></a>] [<a href="http://dx.crossref.org/10.1083/jcb.145.4.899" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>5.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.5">Hemmerich S, Bistrup A, Singer MS, van Zante A, Lee JK, Tsay D, Peters M, Carminati JL, Brennan TJ, Caver-Moore K, Leviten M, Fuentes ME, Ruddle NH, Rosen SD. Sulfation of L-selectin ligands by an HEV-restricted sulfotransferase regulates lymphocyte homing to lymph nodes. <span><span class="ref-journal">Immunity. </span>2001;<span class="ref-vol">15</span>(2):237–47.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/11520459" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 11520459</span></a>] [<a href="http://dx.crossref.org/10.1016/s1074-7613(01)00188-1" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>6.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.6">Hiraoka N, Petryniak B, Nakayama J, Tsuboi S, Suzuki M, Yeh JC, Izawa D, Tanaka T, Miyasaka M, Lowe JB, Fukuda M. A novel, high endothelial venule-specific sulfotransferase expresses 6-sulfo sialyl Lewis(x), an L-selectin ligand displayed by CD34. <span><span class="ref-journal">Immunity. </span>1999;<span class="ref-vol">11</span>(1):79–89.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/10435581" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 10435581</span></a>] [<a href="http://dx.crossref.org/10.1016/s1074-7613(00)80083-7" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>7.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.7">Uchimura K, Gauguet JM, Singer MS, Tsay D, Kannagi R, Muramatsu T, von Andrian UH, Rosen SD. A major class of L-selectin ligands is eliminated in mice deficient in two sulfotransferases expressed in high endothelial venules. <span><span class="ref-journal">Nat Immunol. </span>2005;<span class="ref-vol">6</span>(11):1105–13.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/16227986" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 16227986</span></a>] [<a href="http://dx.crossref.org/10.1038/ni1258" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>8.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.8">Kawashima H, Petryniak B, Hiraoka N, Mitoma J, Huckaby V, Nakayama J, Uchimura K, Kadomatsu K, Muramatsu T, Lowe JB, Fukuda M. <em>N</em>-acetylglucosamine-6-O-sulfotransferases 1 and 2 cooperatively control lymphocyte homing through L-selectin ligand biosynthesis in high endothelial venules. <span><span class="ref-journal">Nat Immunol. </span>2005;<span class="ref-vol">6</span>(11):1096–104.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/16227985" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 16227985</span></a>] [<a href="http://dx.crossref.org/10.1038/ni1259" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>9.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.9">Lee JK, Bhakta S, Rosen SD, Hemmerich S. Cloning and characterization of a mammalian N-acetylglucosamine-6-sulfotransferase that is highly restricted to intestinal tissue. <span><span class="ref-journal">Biochem Biophys Res Commun. </span>1999;<span class="ref-vol">263</span>(2):543–9.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/10491328" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 10491328</span></a>] [<a href="http://dx.crossref.org/10.1006/bbrc.1999.1324" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>10.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.10">Narentuya, Takeda-Uchimura Y, Foyez T, Zhang Z, Akama TO, Yagi H, Kato K, Komatsu Y, Kadomatsu K, Uchimura K. GlcNAc6ST3 is a keratan sulfate sulfotransferase for the protein-tyrosine phosphatase PTPRZ in the adult brain. <span><span class="ref-journal">Sci Rep. </span>2019;<span class="ref-vol">9</span>(1):4387.</span> [<a href="/pmc/articles/PMC6416290/" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pmc">PMC free article<span class="bk_prnt">: PMC6416290</span></a>] [<a href="https://pubmed.ncbi.nlm.nih.gov/30867513" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 30867513</span></a>] [<a href="http://dx.crossref.org/10.1038/s41598-019-40901-2" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>11.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.11">Hoshino H, Foyez T, Ohtake-Niimi S, Takeda-Uchimura Y, Michikawa M, Kadomatsu K, Uchimura K. KSGal6ST is essential for the 6-sulfation of galactose within keratan sulfate in early postnatal brain. <span><span class="ref-journal">J Histochem Cytochem. </span>2014;<span class="ref-vol">62</span>(2):145–56.</span> [<a href="/pmc/articles/PMC3902094/" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pmc">PMC free article<span class="bk_prnt">: PMC3902094</span></a>] [<a href="https://pubmed.ncbi.nlm.nih.gov/24152993" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 24152993</span></a>] [<a href="http://dx.crossref.org/10.1369/0022155413511619" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>12.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.12">Takeda-Uchimura Y, Uchimura K, Sugimura T, Yanagawa Y, Kawasaki T, Komatsu Y, Kadomatsu K. Requirement of keratan sulfate proteoglycan phosphacan with a specific sulfation pattern for critical period plasticity in the visual cortex. Exp Neurol. 2015 274(Pt B):145-55. doi: 10.1016/j.expneurol.2015.08.005. PMID: 26277687. [<a href="https://pubmed.ncbi.nlm.nih.gov/26277687" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 26277687</span></a>] [<a href="http://dx.crossref.org/10.1016/j.expneurol.2015.08.005" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>13.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.13">Hirano K, Ohgomori T, Kobayashi K, Tanaka F, Matsumoto T, Natori T, Matsuyama Y, Uchimura K, Sakamoto K, Takeuchi H, Hirakawa A, Suzumura A, Sobue G, Ishiguro N, Imagama S, Kadomatsu K. Ablation of keratan sulfate accelerates early phase pathogenesis of ALS. <span><span class="ref-journal">PLoS One. </span>2013;<span class="ref-vol">8</span>(6):e66969. </span> [<a href="/pmc/articles/PMC3692529/" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pmc">PMC free article<span class="bk_prnt">: PMC3692529</span></a>] [<a href="https://pubmed.ncbi.nlm.nih.gov/23825599" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 23825599</span></a>] [<a href="http://dx.crossref.org/10.1371/journal.pone.0066969" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>14.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.14">Foyez T, Takeda-Uchimura Y, Ishigaki S. Narentuya, Zhang Z, Sobue G, Kadomatsu K, Uchimura K. Microglial keratan sulfate epitope elicits in central nervous tissues of transgenic model mice and patients with amyotrophic lateral sclerosis. <span><span class="ref-journal">Am J Pathol. </span>2015;<span class="ref-vol">185</span>(11):3053–65.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/26362733" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 26362733</span></a>] [<a href="http://dx.crossref.org/10.1016/j.ajpath.2015.07.016" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>15.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.15">Zhang Z, Takeda-Uchimura Y, Foyez T, Ohtake-Niimi S. Narentuya, Akatsu H, Nishitsuji K, Michikawa M, Wyss-Coray T, Kadomatsu K, Uchimura K. Deficiency of a sulfotransferase for sialic acid-modified glycans mitigates Alzheimer's pathology. <span><span class="ref-journal">Proc Natl Acad Sci U S A. </span>2017;<span class="ref-vol">114</span>(14):E2947–E54.</span> [<a href="/pmc/articles/PMC5389269/" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pmc">PMC free article<span class="bk_prnt">: PMC5389269</span></a>] [<a href="https://pubmed.ncbi.nlm.nih.gov/28320965" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 28320965</span></a>] [<a href="http://dx.crossref.org/10.1073/pnas.1615036114" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>16.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.16">Kitagawa H, Fujita M, Ito N, Sugahara K. Molecular cloning and expression of a novel chondroitin 6-<em>O</em>-sulfotransferase. <span><span class="ref-journal">J Biol Chem. </span>2000;<span class="ref-vol">275</span>(28):21075–80.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/10781596" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 10781596</span></a>] [<a href="http://dx.crossref.org/10.1074/jbc.M002101200" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>17.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.17">Uchimura K, Fasakhany F, Kadomatsu K, Matsukawa T, Yamakawa T, Kurosawa N, Muramatsu T. Diversity of <em>N</em>-acetylglucosamine-6-<em>O</em>-sulfotransferases: molecular cloning of a novel enzyme with different distribution and specificities. <span><span class="ref-journal">Biochem Biophys Res Commun. </span>2000;<span class="ref-vol">274</span>(2):291–6.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/10913333" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 10913333</span></a>] [<a href="http://dx.crossref.org/10.1006/bbrc.2000.3141" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>18.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.18">Bhakta S, Bartes A, Bowman KG, Kao WM, Polsky I, Lee JK, Cook BN, Bruehl RE, Rosen SD, Bertozzi CR, Hemmerich S. Sulfation of <em>N</em>-acetylglucosamine by chondroitin 6-sulfotransferase 2 (GST-5). <span><span class="ref-journal">J Biol Chem. </span>2000;<span class="ref-vol">275</span>(51):40226–34.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/10956661" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 10956661</span></a>] [<a href="http://dx.crossref.org/10.1074/jbc.M006414200" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>19.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.19">Akama TO, Nishida K, Nakayama J, Watanabe H, Ozaki K, Nakamura T, Dota A, Kawasaki S, Inoue Y, Maeda N, Yamamoto S, Fujiwara T, Thonar EJ, Shimomura Y, Kinoshita S, Tanigami A, Fukuda MN. Macular corneal dystrophy type I and type II are caused by distinct mutations in a new sulphotransferase gene. <span><span class="ref-journal">Nat Genet. </span>2000;<span class="ref-vol">26</span>(2):237–41.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/11017086" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 11017086</span></a>] [<a href="http://dx.crossref.org/10.1038/79987" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>20.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.20">Uchimura K, El-Fasakhany FM, Hori M, Hemmerich S, Blink SE, Kansas GS, Kanamori A, Kumamoto K, Kannagi R, Muramatsu T. Specificities of <em>N</em>-acetylglucosamine-6-<em>O</em>-sulfotransferases in relation to L-selectin ligand synthesis and tumor-associated enzyme expression. <span><span class="ref-journal">J Biol Chem. </span>2002;<span class="ref-vol">277</span>(6):3979–84.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/11726653" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 11726653</span></a>] [<a href="http://dx.crossref.org/10.1074/jbc.M106587200" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd><dt>21.</dt><dd><div class="bk_ref" id="g83-assay6st.REF.21">Akama TO, Nakayama J, Nishida K, Hiraoka N, Suzuki M, McAuliffe J, Hindsgaul O, Fukuda M, Fukuda MN. Human corneal GlcNac 6-<em>O</em>-sulfotransferase and mouse intestinal GlcNac 6-<em>O</em>-sulfotransferase both produce keratan sulfate. <span><span class="ref-journal">J Biol Chem. </span>2001;<span class="ref-vol">276</span>(19):16271–8.</span> [<a href="https://pubmed.ncbi.nlm.nih.gov/11278593" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 11278593</span></a>] [<a href="http://dx.crossref.org/10.1074/jbc.M009995200" ref="pagearea=cite-ref&targetsite=external&targetcat=link&targettype=uri">CrossRef</a>]</div></dd></dl></div><h2 id="NBK593984_footnotes">Footnotes</h2><dl class="temp-labeled-list small"><dt></dt><dd><div id="g83-assay6st.FN1"><p class="no_top_margin">The authors declare no competing or financial interests.</p></div></dd></dl><div class="bk_prnt_sctn"><h2>Tables</h2><div class="whole_rhythm bk_prnt_obj bk_first_prnt_obj"><div id="g83-assay6st.T.comparison_of_the_substra" class="table"><h3><span class="label">Table 1: </span></h3><div class="caption"><p>Comparison of the substrate specificity of GlcNAc6ST1, GlcNAc6ST2, GlcNAc6ST3, and GlcNAc6ST4.</p></div><p class="large-table-link" style="display:none"><span class="right"><a href="/books/NBK593984/table/g83-assay6st.T.comparison_of_the_substra/?report=objectonly" target="object">View in own window</a></span></p><div class="large_tbl" id="__g83-assay6st.T.comparison_of_the_substra_lrgtbl__"><table class="no_bottom_margin"><thead><tr><th id="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_1" scope="col" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;"></th><th id="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2" colspan="3" scope="colgroup" rowspan="1" style="text-align:left;vertical-align:top;">Enzyme activity¶</th><th id="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_3" scope="col" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;"></th></tr><tr><th headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_1" id="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_1" scope="col" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Acceptor</th><th headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2" id="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_2" scope="col" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">GlcNAc6ST1</th><th headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2" id="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_3" scope="col" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">GlcNAc6ST2</th><th headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2" id="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_4" scope="col" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">GlcNAc6ST3</th><th headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_3" id="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_5" scope="col" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">GlcNAc6ST4</th></tr></thead><tbody><tr><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_1 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_1" scope="row" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">GlcNAcβ1-6ManOMe</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">20.6 (100)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">6.8 (100)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_4" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">N.D.</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_3 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_5" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">2.8 (100)</td></tr><tr><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_1 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_1" scope="row" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">GlcNAcß1-2Man</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">26.3 (128)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">10.3 (151)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_4" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">N.D.</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_3 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_5" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">2.5 (91)</td></tr><tr><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_1 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_1" scope="row" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">GlcNAcβ1-6[Galβ1-3]GalNAc-pNP (core 2)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">39.3 (191)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">9.9 (145)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_4" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">5.7‡</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_3 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_5" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">0.7 (25)</td></tr><tr><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_1 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_1" scope="row" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">GlcNAcβ1-3GalNAc-pNP (core 3)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">N.D.§</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">12.5 (184)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_4" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">N.D.</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_3 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_5" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">-†</td></tr><tr><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_1 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_1" scope="row" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">GlcNAcβ1-3Galβ1-4GlcNAcβ1-3Galβ1-4GlcNAc</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">4.3‡ (21)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">5.0‡ (73)</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_2 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_4" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">N.D.</td><td headers="hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_1_3 hd_h_g83-assay6st.T.comparison_of_the_substra_1_1_2_5" rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">0.14 (5)</td></tr></tbody></table></div><div><div><dl class="temp-labeled-list small"><dt>¶</dt><dd><div id="g83-assay6st.TF.1.1"><p class="no_margin">The values for GlcNAc6ST1, 2, and 3 were obtained with their protein-A fused versions [pmol/h/ml of medium]. For GlcNAc6ST4, the values were obtained with microsome fractions [pmol/min/mg protein]. The percentage of the activity compared with that of GlcNAcβ1-6ManOMe is also shown. §N.D., less than 0.1 pmol/h/ml of medium. ‡The actually observed radioactivities were around 30,000 cpm, while the assay without the enzyme or with IgG-Sepharose exposed to culture supernatant of mock-transfected cells gave values less than 500 cpm. † -, not tested. Data in previous reports (<a class="bk_pop" href="#g83-assay6st.REF.17">17</a>, <a class="bk_pop" href="#g83-assay6st.REF.20">20</a>) were adopted.</p></div></dd></dl></div></div><div class="permissions"><a href="/books/about/copyright/">Copyright</a> © 2002, © the American Society for Biochemistry and Molecular Biology.</div></div></div></div><div id="bk_toc_contnr"></div></div></div>
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