O-Linked Glycosylation Leads to Decreased Thermal Stability of Interferon Alpha 2b as Measured by Two Orthogonal Techniques
ABSTRACT Purpose Thermal stability is considered an indication of protein fold and conformational stability. We investigate the influence of glycosylation on the thermal stability of interferon alpha 2b (IFN α-2b). Methods Far ultraviolet light circular dichroism spectroscopy (UV CD) and differentia...
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| Veröffentlicht in: | Pharmaceutical research 2011-07, Vol.28 (7), p.1661-1667 |
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| creator | Johnston, Michael James Wilson Frahm, Grant Li, Xuguang Durocher, Yves Hefford, Mary Alice |
| description | ABSTRACT
Purpose
Thermal stability is considered an indication of protein fold and conformational stability. We investigate the influence of glycosylation on the thermal stability of interferon alpha 2b (IFN α-2b).
Methods
Far ultraviolet light circular dichroism spectroscopy (UV CD) and differential scanning calorimetry (DSC) were used to assess the thermal stability of the European Directorate for the Quality of Medicines IFN α-2b reference standards as well as an O-linked glycosylated IFN α-2b produced in human embryonic kidney cells.
Results
Assessment of thermal stability of IFN α-2b and glycosylated IFN α-2b by DSC revealed that non-glycosylated interferon (Tm = 65.7 +/− 0.2°C,
n
= 3) was more thermally stable than the glycosylated variant (Tm = 63.8 C +/− 0.4°C,
n
= 3). These observations were confirmed with far UV CD (Tm IFN α-2b = 65.3 +/− 0.4°C, Tm glycosylated IFN α-2b = 63.6 +/− 0.2°C,
n
= 3). Enzymatic deglycosylation of IFN α-2b resulted in improved thermally stability when assessed with far UV CD and DSC.
Conclusion
We demonstrate that O-linked glycosylation decreases the thermal stability of IFN α-2b compared to a non-glycosylated variant of the protein. |
| doi_str_mv | 10.1007/s11095-011-0402-0 |
| format | Article |
| fullrecord | <record><control><sourceid>proquest_cross</sourceid><recordid>TN_cdi_proquest_miscellaneous_871001267</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>871001267</sourcerecordid><originalsourceid>FETCH-LOGICAL-c508t-1be87a3f0aa95595f0b4f153f37e5b21b20c94eb687dc860e7788c17ccf73bf73</originalsourceid><addsrcrecordid>eNp1kcGKFDEQhoMo7uzqA3iRIMieWqu6k0n6uOzqujAyB0fw1iSZyk6vPZ0x6UYaX940M7ogeChyqO-rKvIz9grhHQKo9wkRalkAYgECygKesAVKVRU1iG9P2QJUKQqtBJ6x85QeAEBjLZ6zsxIrrVDUC_ZrXaza_jtt-W03uZCmzgxt6PmKzDbxIfAbcpFMysBmR3FvOv5lMLbt2mHiwfO7fqDoKWblqjvsDC8tN4l_zsoYs2QnvvkZ-DoOu3Af-qxvyO369sdI6QV75k2X6OXpvWBfP37YXH8qVuvbu-urVeEk6KFAS1qZyoMxtZS19GCFR1n5SpG0JdoSXC3ILrXaOr0EUkprh8o5ryqb64JdHuceYpj3Ds2-TY66zvQUxtTkrwDAcjmTb_4hH8IY89UzBEJkcJkhPEIuhpQi-eYQ272JU4PQzLk0x1yanEsz59JAdl6fBo92T9u_xp8gMvD2BJjkTOej6V2bHjlRSa0kZq48cim3-nuKjxf-f_tvQnClEA</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>870447106</pqid></control><display><type>article</type><title>O-Linked Glycosylation Leads to Decreased Thermal Stability of Interferon Alpha 2b as Measured by Two Orthogonal Techniques</title><source>MEDLINE</source><source>SpringerNature Journals</source><creator>Johnston, Michael James Wilson ; Frahm, Grant ; Li, Xuguang ; Durocher, Yves ; Hefford, Mary Alice</creator><creatorcontrib>Johnston, Michael James Wilson ; Frahm, Grant ; Li, Xuguang ; Durocher, Yves ; Hefford, Mary Alice</creatorcontrib><description>ABSTRACT
Purpose
Thermal stability is considered an indication of protein fold and conformational stability. We investigate the influence of glycosylation on the thermal stability of interferon alpha 2b (IFN α-2b).
Methods
Far ultraviolet light circular dichroism spectroscopy (UV CD) and differential scanning calorimetry (DSC) were used to assess the thermal stability of the European Directorate for the Quality of Medicines IFN α-2b reference standards as well as an O-linked glycosylated IFN α-2b produced in human embryonic kidney cells.
Results
Assessment of thermal stability of IFN α-2b and glycosylated IFN α-2b by DSC revealed that non-glycosylated interferon (Tm = 65.7 +/− 0.2°C,
n
= 3) was more thermally stable than the glycosylated variant (Tm = 63.8 C +/− 0.4°C,
n
= 3). These observations were confirmed with far UV CD (Tm IFN α-2b = 65.3 +/− 0.4°C, Tm glycosylated IFN α-2b = 63.6 +/− 0.2°C,
n
= 3). Enzymatic deglycosylation of IFN α-2b resulted in improved thermally stability when assessed with far UV CD and DSC.
Conclusion
We demonstrate that O-linked glycosylation decreases the thermal stability of IFN α-2b compared to a non-glycosylated variant of the protein.</description><identifier>ISSN: 0724-8741</identifier><identifier>EISSN: 1573-904X</identifier><identifier>DOI: 10.1007/s11095-011-0402-0</identifier><identifier>PMID: 21387149</identifier><identifier>CODEN: PHREEB</identifier><language>eng</language><publisher>Boston: Springer US</publisher><subject>Biochemistry ; Biological and medical sciences ; Biomedical and Life Sciences ; Biomedical Engineering and Bioengineering ; Biomedicine ; Calorimetry, Differential Scanning ; Circular Dichroism ; Differential scanning calorimetry ; Drug Stability ; Electrophoresis, Polyacrylamide Gel ; General pharmacology ; Glycosylation ; Humans ; Interferon ; Interferon-alpha - chemistry ; Medical Law ; Medical sciences ; Pharmaceutical sciences ; Pharmaceutical technology. Pharmaceutical industry ; Pharmacology. Drug treatments ; Pharmacology/Toxicology ; Pharmacy ; Protein Stability ; Research Paper ; Temperature</subject><ispartof>Pharmaceutical research, 2011-07, Vol.28 (7), p.1661-1667</ispartof><rights>Her Majesty the Queen in Right of Canada 2011</rights><rights>2015 INIST-CNRS</rights><rights>Springer Science+Business Media, LLC 2011</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c508t-1be87a3f0aa95595f0b4f153f37e5b21b20c94eb687dc860e7788c17ccf73bf73</citedby><cites>FETCH-LOGICAL-c508t-1be87a3f0aa95595f0b4f153f37e5b21b20c94eb687dc860e7788c17ccf73bf73</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://link.springer.com/content/pdf/10.1007/s11095-011-0402-0$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1007/s11095-011-0402-0$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>314,780,784,27924,27925,41488,42557,51319</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=24358751$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/21387149$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Johnston, Michael James Wilson</creatorcontrib><creatorcontrib>Frahm, Grant</creatorcontrib><creatorcontrib>Li, Xuguang</creatorcontrib><creatorcontrib>Durocher, Yves</creatorcontrib><creatorcontrib>Hefford, Mary Alice</creatorcontrib><title>O-Linked Glycosylation Leads to Decreased Thermal Stability of Interferon Alpha 2b as Measured by Two Orthogonal Techniques</title><title>Pharmaceutical research</title><addtitle>Pharm Res</addtitle><addtitle>Pharm Res</addtitle><description>ABSTRACT
Purpose
Thermal stability is considered an indication of protein fold and conformational stability. We investigate the influence of glycosylation on the thermal stability of interferon alpha 2b (IFN α-2b).
Methods
Far ultraviolet light circular dichroism spectroscopy (UV CD) and differential scanning calorimetry (DSC) were used to assess the thermal stability of the European Directorate for the Quality of Medicines IFN α-2b reference standards as well as an O-linked glycosylated IFN α-2b produced in human embryonic kidney cells.
Results
Assessment of thermal stability of IFN α-2b and glycosylated IFN α-2b by DSC revealed that non-glycosylated interferon (Tm = 65.7 +/− 0.2°C,
n
= 3) was more thermally stable than the glycosylated variant (Tm = 63.8 C +/− 0.4°C,
n
= 3). These observations were confirmed with far UV CD (Tm IFN α-2b = 65.3 +/− 0.4°C, Tm glycosylated IFN α-2b = 63.6 +/− 0.2°C,
n
= 3). Enzymatic deglycosylation of IFN α-2b resulted in improved thermally stability when assessed with far UV CD and DSC.
Conclusion
We demonstrate that O-linked glycosylation decreases the thermal stability of IFN α-2b compared to a non-glycosylated variant of the protein.</description><subject>Biochemistry</subject><subject>Biological and medical sciences</subject><subject>Biomedical and Life Sciences</subject><subject>Biomedical Engineering and Bioengineering</subject><subject>Biomedicine</subject><subject>Calorimetry, Differential Scanning</subject><subject>Circular Dichroism</subject><subject>Differential scanning calorimetry</subject><subject>Drug Stability</subject><subject>Electrophoresis, Polyacrylamide Gel</subject><subject>General pharmacology</subject><subject>Glycosylation</subject><subject>Humans</subject><subject>Interferon</subject><subject>Interferon-alpha - chemistry</subject><subject>Medical Law</subject><subject>Medical sciences</subject><subject>Pharmaceutical sciences</subject><subject>Pharmaceutical technology. Pharmaceutical industry</subject><subject>Pharmacology. Drug treatments</subject><subject>Pharmacology/Toxicology</subject><subject>Pharmacy</subject><subject>Protein Stability</subject><subject>Research Paper</subject><subject>Temperature</subject><issn>0724-8741</issn><issn>1573-904X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2011</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><recordid>eNp1kcGKFDEQhoMo7uzqA3iRIMieWqu6k0n6uOzqujAyB0fw1iSZyk6vPZ0x6UYaX940M7ogeChyqO-rKvIz9grhHQKo9wkRalkAYgECygKesAVKVRU1iG9P2QJUKQqtBJ6x85QeAEBjLZ6zsxIrrVDUC_ZrXaza_jtt-W03uZCmzgxt6PmKzDbxIfAbcpFMysBmR3FvOv5lMLbt2mHiwfO7fqDoKWblqjvsDC8tN4l_zsoYs2QnvvkZ-DoOu3Af-qxvyO369sdI6QV75k2X6OXpvWBfP37YXH8qVuvbu-urVeEk6KFAS1qZyoMxtZS19GCFR1n5SpG0JdoSXC3ILrXaOr0EUkprh8o5ryqb64JdHuceYpj3Ds2-TY66zvQUxtTkrwDAcjmTb_4hH8IY89UzBEJkcJkhPEIuhpQi-eYQ272JU4PQzLk0x1yanEsz59JAdl6fBo92T9u_xp8gMvD2BJjkTOej6V2bHjlRSa0kZq48cim3-nuKjxf-f_tvQnClEA</recordid><startdate>20110701</startdate><enddate>20110701</enddate><creator>Johnston, Michael James Wilson</creator><creator>Frahm, Grant</creator><creator>Li, Xuguang</creator><creator>Durocher, Yves</creator><creator>Hefford, Mary Alice</creator><general>Springer US</general><general>Springer</general><general>Springer Nature B.V</general><scope>IQODW</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>3V.</scope><scope>7RV</scope><scope>7TK</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8AO</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>BENPR</scope><scope>CCPQU</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>K9.</scope><scope>KB0</scope><scope>M0S</scope><scope>M1P</scope><scope>NAPCQ</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>7X8</scope></search><sort><creationdate>20110701</creationdate><title>O-Linked Glycosylation Leads to Decreased Thermal Stability of Interferon Alpha 2b as Measured by Two Orthogonal Techniques</title><author>Johnston, Michael James Wilson ; Frahm, Grant ; Li, Xuguang ; Durocher, Yves ; Hefford, Mary Alice</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c508t-1be87a3f0aa95595f0b4f153f37e5b21b20c94eb687dc860e7788c17ccf73bf73</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2011</creationdate><topic>Biochemistry</topic><topic>Biological and medical sciences</topic><topic>Biomedical and Life Sciences</topic><topic>Biomedical Engineering and Bioengineering</topic><topic>Biomedicine</topic><topic>Calorimetry, Differential Scanning</topic><topic>Circular Dichroism</topic><topic>Differential scanning calorimetry</topic><topic>Drug Stability</topic><topic>Electrophoresis, Polyacrylamide Gel</topic><topic>General pharmacology</topic><topic>Glycosylation</topic><topic>Humans</topic><topic>Interferon</topic><topic>Interferon-alpha - chemistry</topic><topic>Medical Law</topic><topic>Medical sciences</topic><topic>Pharmaceutical sciences</topic><topic>Pharmaceutical technology. Pharmaceutical industry</topic><topic>Pharmacology. Drug treatments</topic><topic>Pharmacology/Toxicology</topic><topic>Pharmacy</topic><topic>Protein Stability</topic><topic>Research Paper</topic><topic>Temperature</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Johnston, Michael James Wilson</creatorcontrib><creatorcontrib>Frahm, Grant</creatorcontrib><creatorcontrib>Li, Xuguang</creatorcontrib><creatorcontrib>Durocher, Yves</creatorcontrib><creatorcontrib>Hefford, Mary Alice</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>Nursing & Allied Health Database</collection><collection>Neurosciences Abstracts</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>ProQuest Pharma Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central</collection><collection>ProQuest One Community College</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>Nursing & Allied Health Database (Alumni Edition)</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Nursing & Allied Health Premium</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>MEDLINE - Academic</collection><jtitle>Pharmaceutical research</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Johnston, Michael James Wilson</au><au>Frahm, Grant</au><au>Li, Xuguang</au><au>Durocher, Yves</au><au>Hefford, Mary Alice</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>O-Linked Glycosylation Leads to Decreased Thermal Stability of Interferon Alpha 2b as Measured by Two Orthogonal Techniques</atitle><jtitle>Pharmaceutical research</jtitle><stitle>Pharm Res</stitle><addtitle>Pharm Res</addtitle><date>2011-07-01</date><risdate>2011</risdate><volume>28</volume><issue>7</issue><spage>1661</spage><epage>1667</epage><pages>1661-1667</pages><issn>0724-8741</issn><eissn>1573-904X</eissn><coden>PHREEB</coden><abstract>ABSTRACT
Purpose
Thermal stability is considered an indication of protein fold and conformational stability. We investigate the influence of glycosylation on the thermal stability of interferon alpha 2b (IFN α-2b).
Methods
Far ultraviolet light circular dichroism spectroscopy (UV CD) and differential scanning calorimetry (DSC) were used to assess the thermal stability of the European Directorate for the Quality of Medicines IFN α-2b reference standards as well as an O-linked glycosylated IFN α-2b produced in human embryonic kidney cells.
Results
Assessment of thermal stability of IFN α-2b and glycosylated IFN α-2b by DSC revealed that non-glycosylated interferon (Tm = 65.7 +/− 0.2°C,
n
= 3) was more thermally stable than the glycosylated variant (Tm = 63.8 C +/− 0.4°C,
n
= 3). These observations were confirmed with far UV CD (Tm IFN α-2b = 65.3 +/− 0.4°C, Tm glycosylated IFN α-2b = 63.6 +/− 0.2°C,
n
= 3). Enzymatic deglycosylation of IFN α-2b resulted in improved thermally stability when assessed with far UV CD and DSC.
Conclusion
We demonstrate that O-linked glycosylation decreases the thermal stability of IFN α-2b compared to a non-glycosylated variant of the protein.</abstract><cop>Boston</cop><pub>Springer US</pub><pmid>21387149</pmid><doi>10.1007/s11095-011-0402-0</doi><tpages>7</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Biochemistry Biological and medical sciences Biomedical and Life Sciences Biomedical Engineering and Bioengineering Biomedicine Calorimetry, Differential Scanning Circular Dichroism Differential scanning calorimetry Drug Stability Electrophoresis, Polyacrylamide Gel General pharmacology Glycosylation Humans Interferon Interferon-alpha - chemistry Medical Law Medical sciences Pharmaceutical sciences Pharmaceutical technology. Pharmaceutical industry Pharmacology. Drug treatments Pharmacology/Toxicology Pharmacy Protein Stability Research Paper Temperature |
| title | O-Linked Glycosylation Leads to Decreased Thermal Stability of Interferon Alpha 2b as Measured by Two Orthogonal Techniques |
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