Plant-Expressed Recombinant Mountain Cedar Allergen Jun a 1 Is Allergenic and Has Limited Pectate Lyase Activity
Background: Mountain cedar (Juniperus ashei) pollen commonly causes a winter time allergic rhinitis in the central USA. Jun a 1 is the dominant allergenic protein, but biologically active recombinant Jun a 1 has not been successfully expressed, despite numerous attempts with several expression syste...
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| Veröffentlicht in: | International archives of allergy and immunology 2010-01, Vol.153 (4), p.347-358 |
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| creator | Liu, Zun Bhattacharyya, Shikha Ning, Bo Midoro-Horiuti, Terumi Czerwinski, Edmund W. Goldblum, Randall M. Mort, Andrew Kearney, Christopher M. |
| description | Background: Mountain cedar (Juniperus ashei) pollen commonly causes a winter time allergic rhinitis in the central USA. Jun a 1 is the dominant allergenic protein, but biologically active recombinant Jun a 1 has not been successfully expressed, despite numerous attempts with several expression systems. Method: Jun a 1 cDNA was inserted into a tobacco mosaic virus vector and transferred to Agrobacterium tumefaciens. Bacteria were syringe-inoculated into leaves of Nicotiana benthamiana (agroinoculation). The interstitial (apoplastic) fluid containing Jun a 1 was isolated. The recombinant protein was analyzed by SDS-PAGE, N-terminal sequencing and MALDI-TOF to confirm identity. Immunogenicity was examined with IgE from allergic patient’s sera, mouse monoclonal anti-Jun a 1 antibodies, IgE-binding inhibition and by degranulation of RBL SX-38 cells sensitized with sera from allergic patients. Pectate lyase activity was assayed by capillary zone electrophoresis and mass spectrometry analysis. Results: Recombinant Jun a 1 was recovered in good quantity (100 µg/g leaf material), was confirmed as Jun a 1, bound IgE from sera from cedar hypersensitive patients and inhibited IgE binding to native Jun a 1. Jun a 1 mutants were created and their pectate lyase activity quantified. For the first time, Jun a 1 pectate lyase activity was demonstrated, which may explain the necrosis seen on host plants, which was similar to that of control plants expressing banana pectate lyase. Conclusions: A means of producing recombinant Jun a 1 is now available for structure/function studies and potentially for diagnostic and therapeutic uses. |
| doi_str_mv | 10.1159/000316345 |
| format | Article |
| fullrecord | <record><control><sourceid>proquest_pubme</sourceid><recordid>TN_cdi_pubmedcentral_primary_oai_pubmedcentral_nih_gov_2945266</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>853474248</sourcerecordid><originalsourceid>FETCH-LOGICAL-c484t-379d8ca8686563e54d30cc11330f5c9ffd43288309e8f9239bfbfd128a7e04a83</originalsourceid><addsrcrecordid>eNpdkU1v1DAQhi1ERUvhwB0hCwkhDgF_JXEuSKtVoUWLqBCcrVlnUlwSZ2s7Ffvv69UuKeVka_z4mbFfQl5w9p7zsvnAGJO8kqp8RE64ErJgrKkf5z3juhBK6mPyNMZrxjKsqyfkWLCybPKtE7K57MGn4uzPJmCM2NLvaMdh7Xyu0q_j5BM4T5fYQqCLvsdwhZ5-mTwFyulFnGvOUvAtPYdIV25wKZsu0SZISFdbiEgXNrlbl7bPyFEHfcTnh_WU_Px09mN5Xqy-fb5YLlaFVVqlQtZNqy3oSldlJbFUrWTWci4l60rbdF2rpNBasgZ11wjZrLt113KhoUamQMtT8nHv3UzrAVuLPgXozSa4AcLWjODMwxPvfpmr8daIRpWiqrLg7UEQxpsJYzKDixb7_F84TtHoUqpaCbVr9fo_8nqcgs-vM5rVleCy5hl6t4dsGGMM2M2jcGZ2KZo5xcy--nf2mfwbWwbeHACIFvougLcu3nNZwiuxE73cc78hhxRm4NDnDiHXrJk</addsrcrecordid><sourcetype>Open Access Repository</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>807621371</pqid></control><display><type>article</type><title>Plant-Expressed Recombinant Mountain Cedar Allergen Jun a 1 Is Allergenic and Has Limited Pectate Lyase Activity</title><source>MEDLINE</source><source>Karger_医学期刊</source><source>Alma/SFX Local Collection</source><creator>Liu, Zun ; Bhattacharyya, Shikha ; Ning, Bo ; Midoro-Horiuti, Terumi ; Czerwinski, Edmund W. ; Goldblum, Randall M. ; Mort, Andrew ; Kearney, Christopher M.</creator><creatorcontrib>Liu, Zun ; Bhattacharyya, Shikha ; Ning, Bo ; Midoro-Horiuti, Terumi ; Czerwinski, Edmund W. ; Goldblum, Randall M. ; Mort, Andrew ; Kearney, Christopher M.</creatorcontrib><description>Background: Mountain cedar (Juniperus ashei) pollen commonly causes a winter time allergic rhinitis in the central USA. Jun a 1 is the dominant allergenic protein, but biologically active recombinant Jun a 1 has not been successfully expressed, despite numerous attempts with several expression systems. Method: Jun a 1 cDNA was inserted into a tobacco mosaic virus vector and transferred to Agrobacterium tumefaciens. Bacteria were syringe-inoculated into leaves of Nicotiana benthamiana (agroinoculation). The interstitial (apoplastic) fluid containing Jun a 1 was isolated. The recombinant protein was analyzed by SDS-PAGE, N-terminal sequencing and MALDI-TOF to confirm identity. Immunogenicity was examined with IgE from allergic patient’s sera, mouse monoclonal anti-Jun a 1 antibodies, IgE-binding inhibition and by degranulation of RBL SX-38 cells sensitized with sera from allergic patients. Pectate lyase activity was assayed by capillary zone electrophoresis and mass spectrometry analysis. Results: Recombinant Jun a 1 was recovered in good quantity (100 µg/g leaf material), was confirmed as Jun a 1, bound IgE from sera from cedar hypersensitive patients and inhibited IgE binding to native Jun a 1. Jun a 1 mutants were created and their pectate lyase activity quantified. For the first time, Jun a 1 pectate lyase activity was demonstrated, which may explain the necrosis seen on host plants, which was similar to that of control plants expressing banana pectate lyase. Conclusions: A means of producing recombinant Jun a 1 is now available for structure/function studies and potentially for diagnostic and therapeutic uses.</description><identifier>ISSN: 1018-2438</identifier><identifier>EISSN: 1423-0097</identifier><identifier>DOI: 10.1159/000316345</identifier><identifier>PMID: 20559000</identifier><language>eng</language><publisher>Basel, Switzerland: Karger</publisher><subject>Agrobacterium tumefaciens ; Agrobacterium tumefaciens - genetics ; Allergens - genetics ; Allergens - immunology ; Allergens - isolation & purification ; Allergens - metabolism ; Allergies ; Animals ; Antigens, Plant ; Basophils - immunology ; Basophils - metabolism ; Basophils - pathology ; Biological and medical sciences ; Cell Degranulation ; Cell Line ; Fundamental and applied biological sciences. Psychology ; Fundamental immunology ; Genetic Vectors - genetics ; Humans ; Immunoglobulin E - metabolism ; Immunopathology ; Juniperus - immunology ; Juniperus ashei ; Medical sciences ; Musa ; Mutagenesis, Site-Directed ; Mutation - genetics ; Nicotiana ; Nicotiana benthamiana ; Original Paper ; Plant Proteins - genetics ; Plant Proteins - immunology ; Plant Proteins - isolation & purification ; Plant Proteins - metabolism ; Pollen ; Polysaccharide-Lyases - metabolism ; Protein Binding ; Rats ; Recombinant Proteins - genetics ; Rhinitis, Allergic, Seasonal - immunology ; Sarcoidosis. Granulomatous diseases of unproved etiology. Connective tissue diseases. Elastic tissue diseases. Vasculitis ; Tobacco mosaic virus ; Tobacco Mosaic Virus - genetics ; Trees</subject><ispartof>International archives of allergy and immunology, 2010-01, Vol.153 (4), p.347-358</ispartof><rights>2010 S. Karger AG, Basel</rights><rights>2015 INIST-CNRS</rights><rights>Copyright © 2010 S. Karger AG, Basel.</rights><rights>Copyright (c) 2010 S. Karger AG, Basel</rights><rights>Copyright © 2010 by S. Karger AG, Basel 2010</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c484t-379d8ca8686563e54d30cc11330f5c9ffd43288309e8f9239bfbfd128a7e04a83</citedby><cites>FETCH-LOGICAL-c484t-379d8ca8686563e54d30cc11330f5c9ffd43288309e8f9239bfbfd128a7e04a83</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>230,314,780,784,885,2427,27915,27916</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=23451625$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/20559000$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Liu, Zun</creatorcontrib><creatorcontrib>Bhattacharyya, Shikha</creatorcontrib><creatorcontrib>Ning, Bo</creatorcontrib><creatorcontrib>Midoro-Horiuti, Terumi</creatorcontrib><creatorcontrib>Czerwinski, Edmund W.</creatorcontrib><creatorcontrib>Goldblum, Randall M.</creatorcontrib><creatorcontrib>Mort, Andrew</creatorcontrib><creatorcontrib>Kearney, Christopher M.</creatorcontrib><title>Plant-Expressed Recombinant Mountain Cedar Allergen Jun a 1 Is Allergenic and Has Limited Pectate Lyase Activity</title><title>International archives of allergy and immunology</title><addtitle>Int Arch Allergy Immunol</addtitle><description>Background: Mountain cedar (Juniperus ashei) pollen commonly causes a winter time allergic rhinitis in the central USA. Jun a 1 is the dominant allergenic protein, but biologically active recombinant Jun a 1 has not been successfully expressed, despite numerous attempts with several expression systems. Method: Jun a 1 cDNA was inserted into a tobacco mosaic virus vector and transferred to Agrobacterium tumefaciens. Bacteria were syringe-inoculated into leaves of Nicotiana benthamiana (agroinoculation). The interstitial (apoplastic) fluid containing Jun a 1 was isolated. The recombinant protein was analyzed by SDS-PAGE, N-terminal sequencing and MALDI-TOF to confirm identity. Immunogenicity was examined with IgE from allergic patient’s sera, mouse monoclonal anti-Jun a 1 antibodies, IgE-binding inhibition and by degranulation of RBL SX-38 cells sensitized with sera from allergic patients. Pectate lyase activity was assayed by capillary zone electrophoresis and mass spectrometry analysis. Results: Recombinant Jun a 1 was recovered in good quantity (100 µg/g leaf material), was confirmed as Jun a 1, bound IgE from sera from cedar hypersensitive patients and inhibited IgE binding to native Jun a 1. Jun a 1 mutants were created and their pectate lyase activity quantified. For the first time, Jun a 1 pectate lyase activity was demonstrated, which may explain the necrosis seen on host plants, which was similar to that of control plants expressing banana pectate lyase. Conclusions: A means of producing recombinant Jun a 1 is now available for structure/function studies and potentially for diagnostic and therapeutic uses.</description><subject>Agrobacterium tumefaciens</subject><subject>Agrobacterium tumefaciens - genetics</subject><subject>Allergens - genetics</subject><subject>Allergens - immunology</subject><subject>Allergens - isolation & purification</subject><subject>Allergens - metabolism</subject><subject>Allergies</subject><subject>Animals</subject><subject>Antigens, Plant</subject><subject>Basophils - immunology</subject><subject>Basophils - metabolism</subject><subject>Basophils - pathology</subject><subject>Biological and medical sciences</subject><subject>Cell Degranulation</subject><subject>Cell Line</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Fundamental immunology</subject><subject>Genetic Vectors - genetics</subject><subject>Humans</subject><subject>Immunoglobulin E - metabolism</subject><subject>Immunopathology</subject><subject>Juniperus - immunology</subject><subject>Juniperus ashei</subject><subject>Medical sciences</subject><subject>Musa</subject><subject>Mutagenesis, Site-Directed</subject><subject>Mutation - genetics</subject><subject>Nicotiana</subject><subject>Nicotiana benthamiana</subject><subject>Original Paper</subject><subject>Plant Proteins - genetics</subject><subject>Plant Proteins - immunology</subject><subject>Plant Proteins - isolation & purification</subject><subject>Plant Proteins - metabolism</subject><subject>Pollen</subject><subject>Polysaccharide-Lyases - metabolism</subject><subject>Protein Binding</subject><subject>Rats</subject><subject>Recombinant Proteins - genetics</subject><subject>Rhinitis, Allergic, Seasonal - immunology</subject><subject>Sarcoidosis. Granulomatous diseases of unproved etiology. Connective tissue diseases. Elastic tissue diseases. Vasculitis</subject><subject>Tobacco mosaic virus</subject><subject>Tobacco Mosaic Virus - genetics</subject><subject>Trees</subject><issn>1018-2438</issn><issn>1423-0097</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2010</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>8G5</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><sourceid>GUQSH</sourceid><sourceid>M2O</sourceid><recordid>eNpdkU1v1DAQhi1ERUvhwB0hCwkhDgF_JXEuSKtVoUWLqBCcrVlnUlwSZ2s7Ffvv69UuKeVka_z4mbFfQl5w9p7zsvnAGJO8kqp8RE64ErJgrKkf5z3juhBK6mPyNMZrxjKsqyfkWLCybPKtE7K57MGn4uzPJmCM2NLvaMdh7Xyu0q_j5BM4T5fYQqCLvsdwhZ5-mTwFyulFnGvOUvAtPYdIV25wKZsu0SZISFdbiEgXNrlbl7bPyFEHfcTnh_WU_Px09mN5Xqy-fb5YLlaFVVqlQtZNqy3oSldlJbFUrWTWci4l60rbdF2rpNBasgZ11wjZrLt113KhoUamQMtT8nHv3UzrAVuLPgXozSa4AcLWjODMwxPvfpmr8daIRpWiqrLg7UEQxpsJYzKDixb7_F84TtHoUqpaCbVr9fo_8nqcgs-vM5rVleCy5hl6t4dsGGMM2M2jcGZ2KZo5xcy--nf2mfwbWwbeHACIFvougLcu3nNZwiuxE73cc78hhxRm4NDnDiHXrJk</recordid><startdate>20100101</startdate><enddate>20100101</enddate><creator>Liu, Zun</creator><creator>Bhattacharyya, Shikha</creator><creator>Ning, Bo</creator><creator>Midoro-Horiuti, Terumi</creator><creator>Czerwinski, Edmund W.</creator><creator>Goldblum, Randall M.</creator><creator>Mort, Andrew</creator><creator>Kearney, Christopher M.</creator><general>Karger</general><general>S. 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Psychology</topic><topic>Fundamental immunology</topic><topic>Genetic Vectors - genetics</topic><topic>Humans</topic><topic>Immunoglobulin E - metabolism</topic><topic>Immunopathology</topic><topic>Juniperus - immunology</topic><topic>Juniperus ashei</topic><topic>Medical sciences</topic><topic>Musa</topic><topic>Mutagenesis, Site-Directed</topic><topic>Mutation - genetics</topic><topic>Nicotiana</topic><topic>Nicotiana benthamiana</topic><topic>Original Paper</topic><topic>Plant Proteins - genetics</topic><topic>Plant Proteins - immunology</topic><topic>Plant Proteins - isolation & purification</topic><topic>Plant Proteins - metabolism</topic><topic>Pollen</topic><topic>Polysaccharide-Lyases - metabolism</topic><topic>Protein Binding</topic><topic>Rats</topic><topic>Recombinant Proteins - genetics</topic><topic>Rhinitis, Allergic, Seasonal - immunology</topic><topic>Sarcoidosis. Granulomatous diseases of unproved etiology. Connective tissue diseases. Elastic tissue diseases. Vasculitis</topic><topic>Tobacco mosaic virus</topic><topic>Tobacco Mosaic Virus - genetics</topic><topic>Trees</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Liu, Zun</creatorcontrib><creatorcontrib>Bhattacharyya, Shikha</creatorcontrib><creatorcontrib>Ning, Bo</creatorcontrib><creatorcontrib>Midoro-Horiuti, Terumi</creatorcontrib><creatorcontrib>Czerwinski, Edmund W.</creatorcontrib><creatorcontrib>Goldblum, Randall M.</creatorcontrib><creatorcontrib>Mort, Andrew</creatorcontrib><creatorcontrib>Kearney, Christopher M.</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>ProQuest Central (Corporate)</collection><collection>Immunology Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>ProQuest Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>ProQuest Pharma Collection</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Natural Science Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>Research Library (Alumni Edition)</collection><collection>ProQuest Central (Alumni)</collection><collection>ProQuest Central</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Natural Science Collection</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Central Student</collection><collection>Research Library Prep</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>SciTech Premium Collection (Proquest) (PQ_SDU_P3)</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>Biological Sciences</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>PML(ProQuest Medical Library)</collection><collection>ProQuest research library</collection><collection>Biological Science Database</collection><collection>Research Library (Corporate)</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central China</collection><collection>ProQuest Central Basic</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>International archives of allergy and immunology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Liu, Zun</au><au>Bhattacharyya, Shikha</au><au>Ning, Bo</au><au>Midoro-Horiuti, Terumi</au><au>Czerwinski, Edmund W.</au><au>Goldblum, Randall M.</au><au>Mort, Andrew</au><au>Kearney, Christopher M.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Plant-Expressed Recombinant Mountain Cedar Allergen Jun a 1 Is Allergenic and Has Limited Pectate Lyase Activity</atitle><jtitle>International archives of allergy and immunology</jtitle><addtitle>Int Arch Allergy Immunol</addtitle><date>2010-01-01</date><risdate>2010</risdate><volume>153</volume><issue>4</issue><spage>347</spage><epage>358</epage><pages>347-358</pages><issn>1018-2438</issn><eissn>1423-0097</eissn><abstract>Background: Mountain cedar (Juniperus ashei) pollen commonly causes a winter time allergic rhinitis in the central USA. Jun a 1 is the dominant allergenic protein, but biologically active recombinant Jun a 1 has not been successfully expressed, despite numerous attempts with several expression systems. Method: Jun a 1 cDNA was inserted into a tobacco mosaic virus vector and transferred to Agrobacterium tumefaciens. Bacteria were syringe-inoculated into leaves of Nicotiana benthamiana (agroinoculation). The interstitial (apoplastic) fluid containing Jun a 1 was isolated. The recombinant protein was analyzed by SDS-PAGE, N-terminal sequencing and MALDI-TOF to confirm identity. Immunogenicity was examined with IgE from allergic patient’s sera, mouse monoclonal anti-Jun a 1 antibodies, IgE-binding inhibition and by degranulation of RBL SX-38 cells sensitized with sera from allergic patients. Pectate lyase activity was assayed by capillary zone electrophoresis and mass spectrometry analysis. Results: Recombinant Jun a 1 was recovered in good quantity (100 µg/g leaf material), was confirmed as Jun a 1, bound IgE from sera from cedar hypersensitive patients and inhibited IgE binding to native Jun a 1. Jun a 1 mutants were created and their pectate lyase activity quantified. For the first time, Jun a 1 pectate lyase activity was demonstrated, which may explain the necrosis seen on host plants, which was similar to that of control plants expressing banana pectate lyase. Conclusions: A means of producing recombinant Jun a 1 is now available for structure/function studies and potentially for diagnostic and therapeutic uses.</abstract><cop>Basel, Switzerland</cop><pub>Karger</pub><pmid>20559000</pmid><doi>10.1159/000316345</doi><tpages>12</tpages><oa>free_for_read</oa></addata></record> |
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| language | eng |
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| source | MEDLINE; Karger_医学期刊; Alma/SFX Local Collection |
| subjects | Agrobacterium tumefaciens Agrobacterium tumefaciens - genetics Allergens - genetics Allergens - immunology Allergens - isolation & purification Allergens - metabolism Allergies Animals Antigens, Plant Basophils - immunology Basophils - metabolism Basophils - pathology Biological and medical sciences Cell Degranulation Cell Line Fundamental and applied biological sciences. Psychology Fundamental immunology Genetic Vectors - genetics Humans Immunoglobulin E - metabolism Immunopathology Juniperus - immunology Juniperus ashei Medical sciences Musa Mutagenesis, Site-Directed Mutation - genetics Nicotiana Nicotiana benthamiana Original Paper Plant Proteins - genetics Plant Proteins - immunology Plant Proteins - isolation & purification Plant Proteins - metabolism Pollen Polysaccharide-Lyases - metabolism Protein Binding Rats Recombinant Proteins - genetics Rhinitis, Allergic, Seasonal - immunology Sarcoidosis. Granulomatous diseases of unproved etiology. Connective tissue diseases. Elastic tissue diseases. Vasculitis Tobacco mosaic virus Tobacco Mosaic Virus - genetics Trees |
| title | Plant-Expressed Recombinant Mountain Cedar Allergen Jun a 1 Is Allergenic and Has Limited Pectate Lyase Activity |
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