N-glycan structures of human transferrin produced by Lymantria dispar (gypsy moth) cells using the LdMNPV expression system
N-glycan structures of recombinant human serum transferrin (hTf) expressed by Lymantria dispar (gypsy moth) 652Y cells were determined. The gene encoding hTf was incorporated into a Lymantria dispar nucleopolyhedrovirus (LdMNPV) under the control of the polyhedrin promoter. This virus was then used...
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| Veröffentlicht in: | Glycobiology (Oxford) 2003-07, Vol.13 (7), p.539-548 |
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| creator | Choi, One Tomiya, Noboru Kim, Jung H. Slavicek, James M. Betenbaugh, Michael J. Lee, Yuan C. |
| description | N-glycan structures of recombinant human serum transferrin (hTf) expressed by Lymantria dispar (gypsy moth) 652Y cells were determined. The gene encoding hTf was incorporated into a Lymantria dispar nucleopolyhedrovirus (LdMNPV) under the control of the polyhedrin promoter. This virus was then used to infect Ld652Y cells, and the recombinant protein was harvested at 120 h postinfection. N-glycans were released from the purified recombinant human serum transferrin and derivatized with 2-aminopyridine; the glycan structures were analyzed by a two-dimensional HPLC and MALDI-TOF MS. Structures of 11 glycans (88.8% of total N-glycans) were elucidated. The glycan analysis revealed that the most abundant glycans were Man1–3(±Fucα6)GlcNAc2 (75.5%) and GlcNAcMan3(±Fucα6)GlcNAc2 (7.4%). There was only ∼6% of high-mannose type glycans identified. Nearly half (49.8%) of the total N-glycans contained α(1,6)-fucosylation on the Asn-linked GlcNAc residue. However α(1,3)-fucosylation on the same GlcNAc, often found in N-glycans produced by other insects and insect cells, was not detected. Inclusion of fetal bovine serum in culture media had little effect on the N-glycan structures of the recombinant human serum transferrin obtained. |
| doi_str_mv | 10.1093/glycob/cwg071 |
| format | Article |
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The gene encoding hTf was incorporated into a Lymantria dispar nucleopolyhedrovirus (LdMNPV) under the control of the polyhedrin promoter. This virus was then used to infect Ld652Y cells, and the recombinant protein was harvested at 120 h postinfection. N-glycans were released from the purified recombinant human serum transferrin and derivatized with 2-aminopyridine; the glycan structures were analyzed by a two-dimensional HPLC and MALDI-TOF MS. Structures of 11 glycans (88.8% of total N-glycans) were elucidated. The glycan analysis revealed that the most abundant glycans were Man1–3(±Fucα6)GlcNAc2 (75.5%) and GlcNAcMan3(±Fucα6)GlcNAc2 (7.4%). There was only ∼6% of high-mannose type glycans identified. Nearly half (49.8%) of the total N-glycans contained α(1,6)-fucosylation on the Asn-linked GlcNAc residue. However α(1,3)-fucosylation on the same GlcNAc, often found in N-glycans produced by other insects and insect cells, was not detected. Inclusion of fetal bovine serum in culture media had little effect on the N-glycan structures of the recombinant human serum transferrin obtained.</description><identifier>ISSN: 0959-6658</identifier><identifier>EISSN: 1460-2423</identifier><identifier>DOI: 10.1093/glycob/cwg071</identifier><identifier>PMID: 12672704</identifier><language>eng</language><publisher>England: Oxford University Press</publisher><subject>AcMNPV ; Animals ; Autographa californica multicapsid nucleopolyhedrovirus ; Baculovirus ; Carbohydrate Conformation ; Carbohydrate Sequence ; Culture Media ; FBS ; fetal bovine serum ; Gene Expression ; glucose unit ; Gypsy moth ; high-performance anion-exchange chromatography with pulsed amperometric detection ; high-performance liquid chromatography ; HPAEC-PAD ; HPLC ; hTf ; human transferrin ; Humans ; Insect cells ; LdMNPV ; Lepidoptera - genetics ; Lymantria dispar ; Lymantria dispar multicapsid nucleopolyhedrovirus ; Lymantria dispar nucleopolyhedrovirus ; MALDI-TOF ; mass spectrometry ; matrix-assisted laser desorption/ionization time of flight ; Molecular Sequence Data ; N-glycan ; Nuclear polyhedrosis virus ; Polysaccharides - biosynthesis ; Polysaccharides - chemistry ; pyridylamino ; Recombinant Proteins - biosynthesis ; Recombinant Proteins - chemistry ; Recombinant Proteins - genetics ; Recombinant Proteins - isolation & purification ; SDS–PAGE ; sodium dodecyl sulfate–polyacrylamide gel electrophoresis ; TCID50 ; tissue culture infectious dose ; Transferrin - biosynthesis ; Transferrin - chemistry ; Transferrin - genetics ; Transferrin - isolation & purification</subject><ispartof>Glycobiology (Oxford), 2003-07, Vol.13 (7), p.539-548</ispartof><rights>Copyright Oxford University Press(England) Jul 2003</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c490t-ca7f4572c77a8fd04392194230d85595f37060455c75219d120a49f6f75d1d2f3</citedby><cites>FETCH-LOGICAL-c490t-ca7f4572c77a8fd04392194230d85595f37060455c75219d120a49f6f75d1d2f3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/12672704$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Choi, One</creatorcontrib><creatorcontrib>Tomiya, Noboru</creatorcontrib><creatorcontrib>Kim, Jung H.</creatorcontrib><creatorcontrib>Slavicek, James M.</creatorcontrib><creatorcontrib>Betenbaugh, Michael J.</creatorcontrib><creatorcontrib>Lee, Yuan C.</creatorcontrib><title>N-glycan structures of human transferrin produced by Lymantria dispar (gypsy moth) cells using the LdMNPV expression system</title><title>Glycobiology (Oxford)</title><addtitle>Glycobiology</addtitle><description>N-glycan structures of recombinant human serum transferrin (hTf) expressed by Lymantria dispar (gypsy moth) 652Y cells were determined. The gene encoding hTf was incorporated into a Lymantria dispar nucleopolyhedrovirus (LdMNPV) under the control of the polyhedrin promoter. This virus was then used to infect Ld652Y cells, and the recombinant protein was harvested at 120 h postinfection. N-glycans were released from the purified recombinant human serum transferrin and derivatized with 2-aminopyridine; the glycan structures were analyzed by a two-dimensional HPLC and MALDI-TOF MS. Structures of 11 glycans (88.8% of total N-glycans) were elucidated. The glycan analysis revealed that the most abundant glycans were Man1–3(±Fucα6)GlcNAc2 (75.5%) and GlcNAcMan3(±Fucα6)GlcNAc2 (7.4%). There was only ∼6% of high-mannose type glycans identified. Nearly half (49.8%) of the total N-glycans contained α(1,6)-fucosylation on the Asn-linked GlcNAc residue. However α(1,3)-fucosylation on the same GlcNAc, often found in N-glycans produced by other insects and insect cells, was not detected. Inclusion of fetal bovine serum in culture media had little effect on the N-glycan structures of the recombinant human serum transferrin obtained.</description><subject>AcMNPV</subject><subject>Animals</subject><subject>Autographa californica multicapsid nucleopolyhedrovirus</subject><subject>Baculovirus</subject><subject>Carbohydrate Conformation</subject><subject>Carbohydrate Sequence</subject><subject>Culture Media</subject><subject>FBS</subject><subject>fetal bovine serum</subject><subject>Gene Expression</subject><subject>glucose unit</subject><subject>Gypsy moth</subject><subject>high-performance anion-exchange chromatography with pulsed amperometric detection</subject><subject>high-performance liquid chromatography</subject><subject>HPAEC-PAD</subject><subject>HPLC</subject><subject>hTf</subject><subject>human transferrin</subject><subject>Humans</subject><subject>Insect cells</subject><subject>LdMNPV</subject><subject>Lepidoptera - genetics</subject><subject>Lymantria dispar</subject><subject>Lymantria dispar multicapsid nucleopolyhedrovirus</subject><subject>Lymantria dispar nucleopolyhedrovirus</subject><subject>MALDI-TOF</subject><subject>mass spectrometry</subject><subject>matrix-assisted laser desorption/ionization time of flight</subject><subject>Molecular Sequence Data</subject><subject>N-glycan</subject><subject>Nuclear polyhedrosis virus</subject><subject>Polysaccharides - biosynthesis</subject><subject>Polysaccharides - chemistry</subject><subject>pyridylamino</subject><subject>Recombinant Proteins - biosynthesis</subject><subject>Recombinant Proteins - chemistry</subject><subject>Recombinant Proteins - genetics</subject><subject>Recombinant Proteins - isolation & purification</subject><subject>SDS–PAGE</subject><subject>sodium dodecyl sulfate–polyacrylamide gel electrophoresis</subject><subject>TCID50</subject><subject>tissue culture infectious dose</subject><subject>Transferrin - biosynthesis</subject><subject>Transferrin - chemistry</subject><subject>Transferrin - genetics</subject><subject>Transferrin - isolation & purification</subject><issn>0959-6658</issn><issn>1460-2423</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2003</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkUtrVDEYhoNY7LS6dCvBhdTFaXM9OVmWoq0yrQNekG5CJpeZU-dczAV78M-bYYYW3HQVyPfwJO_3AvAao1OMJD1bbSYzLM_MnxUS-BmYYVajijBCn4MZklxWdc2bQ3AU4x1CuMYNfwEOMakFEYjNwN-bamvQPYwpZJNycBEOHq5zV-5S0H30LoS2h2MYbDbOwuUE51OZptBqaNs46gBPVtMYJ9gNaf0eGrfZRJhj269gWjs4t9c3ix_Q3Y9FHtuhvDXF5LqX4MDrTXSv9ucx-P7xw7eLq2r-5fLTxfm8MkyiVBktPOOCGCF04y1iVBIsS0JkG84l91SgGjHOjeBlYDFBmklfe8EttsTTY_Bu5y0RfmcXk-rauP2k7t2QoxKUNkXIngSxbERDuSjg2__AuyGHvoRQBCPKEKKyQNUOMmGIMTivxtB2OkwKI7XtTu26U7vuCv9mL83LztlHel_Wo7At27t_mOvwS9WCCq6uft6q26-L67q5XKjP9B8CnqZV</recordid><startdate>20030701</startdate><enddate>20030701</enddate><creator>Choi, One</creator><creator>Tomiya, Noboru</creator><creator>Kim, Jung H.</creator><creator>Slavicek, James M.</creator><creator>Betenbaugh, Michael J.</creator><creator>Lee, Yuan C.</creator><general>Oxford University Press</general><general>Oxford Publishing Limited (England)</general><scope>BSCLL</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QL</scope><scope>7QO</scope><scope>7TK</scope><scope>7U9</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>H94</scope><scope>K9.</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope><scope>7SS</scope><scope>7X8</scope></search><sort><creationdate>20030701</creationdate><title>N-glycan structures of human transferrin produced by Lymantria dispar (gypsy moth) cells using the LdMNPV expression system</title><author>Choi, One ; Tomiya, Noboru ; Kim, Jung H. ; Slavicek, James M. ; Betenbaugh, Michael J. ; Lee, Yuan C.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c490t-ca7f4572c77a8fd04392194230d85595f37060455c75219d120a49f6f75d1d2f3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2003</creationdate><topic>AcMNPV</topic><topic>Animals</topic><topic>Autographa californica multicapsid nucleopolyhedrovirus</topic><topic>Baculovirus</topic><topic>Carbohydrate Conformation</topic><topic>Carbohydrate Sequence</topic><topic>Culture Media</topic><topic>FBS</topic><topic>fetal bovine serum</topic><topic>Gene Expression</topic><topic>glucose unit</topic><topic>Gypsy moth</topic><topic>high-performance anion-exchange chromatography with pulsed amperometric detection</topic><topic>high-performance liquid chromatography</topic><topic>HPAEC-PAD</topic><topic>HPLC</topic><topic>hTf</topic><topic>human transferrin</topic><topic>Humans</topic><topic>Insect cells</topic><topic>LdMNPV</topic><topic>Lepidoptera - genetics</topic><topic>Lymantria dispar</topic><topic>Lymantria dispar multicapsid nucleopolyhedrovirus</topic><topic>Lymantria dispar nucleopolyhedrovirus</topic><topic>MALDI-TOF</topic><topic>mass spectrometry</topic><topic>matrix-assisted laser desorption/ionization time of flight</topic><topic>Molecular Sequence Data</topic><topic>N-glycan</topic><topic>Nuclear polyhedrosis virus</topic><topic>Polysaccharides - biosynthesis</topic><topic>Polysaccharides - chemistry</topic><topic>pyridylamino</topic><topic>Recombinant Proteins - biosynthesis</topic><topic>Recombinant Proteins - chemistry</topic><topic>Recombinant Proteins - genetics</topic><topic>Recombinant Proteins - isolation & purification</topic><topic>SDS–PAGE</topic><topic>sodium dodecyl sulfate–polyacrylamide gel electrophoresis</topic><topic>TCID50</topic><topic>tissue culture infectious dose</topic><topic>Transferrin - biosynthesis</topic><topic>Transferrin - chemistry</topic><topic>Transferrin - genetics</topic><topic>Transferrin - isolation & purification</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Choi, One</creatorcontrib><creatorcontrib>Tomiya, Noboru</creatorcontrib><creatorcontrib>Kim, Jung H.</creatorcontrib><creatorcontrib>Slavicek, James M.</creatorcontrib><creatorcontrib>Betenbaugh, Michael J.</creatorcontrib><creatorcontrib>Lee, Yuan C.</creatorcontrib><collection>Istex</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Biotechnology Research Abstracts</collection><collection>Neurosciences Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>Entomology Abstracts (Full archive)</collection><collection>MEDLINE - Academic</collection><jtitle>Glycobiology (Oxford)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Choi, One</au><au>Tomiya, Noboru</au><au>Kim, Jung H.</au><au>Slavicek, James M.</au><au>Betenbaugh, Michael J.</au><au>Lee, Yuan C.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>N-glycan structures of human transferrin produced by Lymantria dispar (gypsy moth) cells using the LdMNPV expression system</atitle><jtitle>Glycobiology (Oxford)</jtitle><addtitle>Glycobiology</addtitle><date>2003-07-01</date><risdate>2003</risdate><volume>13</volume><issue>7</issue><spage>539</spage><epage>548</epage><pages>539-548</pages><issn>0959-6658</issn><eissn>1460-2423</eissn><abstract>N-glycan structures of recombinant human serum transferrin (hTf) expressed by Lymantria dispar (gypsy moth) 652Y cells were determined. The gene encoding hTf was incorporated into a Lymantria dispar nucleopolyhedrovirus (LdMNPV) under the control of the polyhedrin promoter. This virus was then used to infect Ld652Y cells, and the recombinant protein was harvested at 120 h postinfection. N-glycans were released from the purified recombinant human serum transferrin and derivatized with 2-aminopyridine; the glycan structures were analyzed by a two-dimensional HPLC and MALDI-TOF MS. Structures of 11 glycans (88.8% of total N-glycans) were elucidated. The glycan analysis revealed that the most abundant glycans were Man1–3(±Fucα6)GlcNAc2 (75.5%) and GlcNAcMan3(±Fucα6)GlcNAc2 (7.4%). There was only ∼6% of high-mannose type glycans identified. Nearly half (49.8%) of the total N-glycans contained α(1,6)-fucosylation on the Asn-linked GlcNAc residue. However α(1,3)-fucosylation on the same GlcNAc, often found in N-glycans produced by other insects and insect cells, was not detected. Inclusion of fetal bovine serum in culture media had little effect on the N-glycan structures of the recombinant human serum transferrin obtained.</abstract><cop>England</cop><pub>Oxford University Press</pub><pmid>12672704</pmid><doi>10.1093/glycob/cwg071</doi><tpages>10</tpages><oa>free_for_read</oa></addata></record> |
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| source | MEDLINE; Oxford University Press Journals All Titles (1996-Current); EZB-FREE-00999 freely available EZB journals; Alma/SFX Local Collection |
| subjects | AcMNPV Animals Autographa californica multicapsid nucleopolyhedrovirus Baculovirus Carbohydrate Conformation Carbohydrate Sequence Culture Media FBS fetal bovine serum Gene Expression glucose unit Gypsy moth high-performance anion-exchange chromatography with pulsed amperometric detection high-performance liquid chromatography HPAEC-PAD HPLC hTf human transferrin Humans Insect cells LdMNPV Lepidoptera - genetics Lymantria dispar Lymantria dispar multicapsid nucleopolyhedrovirus Lymantria dispar nucleopolyhedrovirus MALDI-TOF mass spectrometry matrix-assisted laser desorption/ionization time of flight Molecular Sequence Data N-glycan Nuclear polyhedrosis virus Polysaccharides - biosynthesis Polysaccharides - chemistry pyridylamino Recombinant Proteins - biosynthesis Recombinant Proteins - chemistry Recombinant Proteins - genetics Recombinant Proteins - isolation & purification SDS–PAGE sodium dodecyl sulfate–polyacrylamide gel electrophoresis TCID50 tissue culture infectious dose Transferrin - biosynthesis Transferrin - chemistry Transferrin - genetics Transferrin - isolation & purification |
| title | N-glycan structures of human transferrin produced by Lymantria dispar (gypsy moth) cells using the LdMNPV expression system |
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