Identification and measurement of isoaspartic acid formation in the complementarity determining region of a fully human monoclonal antibody
Isomerization plays a key role in protein degradation. This isomerization is often difficult to detect by many protein characterization methods such as SDS-PAGE, SEC, and IEF. This work shows the identification of an isomerized aspartic acid residue in the CDR2 of the heavy chain of a fully human mo...
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| Veröffentlicht in: | Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Analytical technologies in the biomedical and life sciences, 2009-11, Vol.877 (30), p.3841-3849 |
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| container_title | Journal of chromatography. B, Analytical technologies in the biomedical and life sciences |
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| creator | Dick, Lawrence W. Qiu, Difei Cheng, Kuang-Chuan |
| description | Isomerization plays a key role in protein degradation. This isomerization is often difficult to detect by many protein characterization methods such as SDS-PAGE, SEC, and IEF. This work shows the identification of an isomerized aspartic acid residue in the CDR2 of the heavy chain of a fully human monoclonal antibody. This isoaspartic acid increases significantly with storage at 2–8
°C. Hydrophobic interaction chromatography was utilized to separate the isoaspartic variant in the intact state. Mass spectrometry including peptide mapping was employed to identify and confirm the exact location of the modification. Since this modification occurs in the complementarity determining region (CDR) it was found that binding is reduced. Therefore, three different analytical methods for regular analysis of this isomerization are evaluated. These methods include peptide mapping by LC–MS, HIC, and a protein isoaspartate methyltransferase assay. It was determined that HIC is the best method to regularly assay the level of isomerization in this monoclonal antibody. |
| doi_str_mv | 10.1016/j.jchromb.2009.09.031 |
| format | Article |
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°C. Hydrophobic interaction chromatography was utilized to separate the isoaspartic variant in the intact state. Mass spectrometry including peptide mapping was employed to identify and confirm the exact location of the modification. Since this modification occurs in the complementarity determining region (CDR) it was found that binding is reduced. Therefore, three different analytical methods for regular analysis of this isomerization are evaluated. These methods include peptide mapping by LC–MS, HIC, and a protein isoaspartate methyltransferase assay. It was determined that HIC is the best method to regularly assay the level of isomerization in this monoclonal antibody.</description><identifier>ISSN: 1570-0232</identifier><identifier>EISSN: 1873-376X</identifier><identifier>DOI: 10.1016/j.jchromb.2009.09.031</identifier><identifier>PMID: 19819766</identifier><language>eng</language><publisher>Amsterdam: Elsevier B.V</publisher><subject>Analysis ; Analytical, structural and metabolic biochemistry ; Antibodies, Monoclonal - chemistry ; Biological and medical sciences ; Chromatography, Affinity - methods ; Complementarity Determining Regions - chemistry ; Fundamental and applied biological sciences. Psychology ; General pharmacology ; Humans ; Hydrophobic interaction chromatography ; Isoaspartic Acid - chemistry ; Isomerism ; Isomerization ; Mass Spectrometry - methods ; Medical sciences ; Monoclonal antibody ; Peptide mapping ; Peptide Mapping - methods ; Pharmacology. Drug treatments ; PIMT assay</subject><ispartof>Journal of chromatography. B, Analytical technologies in the biomedical and life sciences, 2009-11, Vol.877 (30), p.3841-3849</ispartof><rights>2009 Elsevier B.V.</rights><rights>2015 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c489t-3cd19c0b48352ad76bb0570962faf1f5078550a58e160fd2f27ec70ae003e8a63</citedby><cites>FETCH-LOGICAL-c489t-3cd19c0b48352ad76bb0570962faf1f5078550a58e160fd2f27ec70ae003e8a63</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/j.jchromb.2009.09.031$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,780,784,3550,27924,27925,45995</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=22161999$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/19819766$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Dick, Lawrence W.</creatorcontrib><creatorcontrib>Qiu, Difei</creatorcontrib><creatorcontrib>Cheng, Kuang-Chuan</creatorcontrib><title>Identification and measurement of isoaspartic acid formation in the complementarity determining region of a fully human monoclonal antibody</title><title>Journal of chromatography. B, Analytical technologies in the biomedical and life sciences</title><addtitle>J Chromatogr B Analyt Technol Biomed Life Sci</addtitle><description>Isomerization plays a key role in protein degradation. This isomerization is often difficult to detect by many protein characterization methods such as SDS-PAGE, SEC, and IEF. This work shows the identification of an isomerized aspartic acid residue in the CDR2 of the heavy chain of a fully human monoclonal antibody. This isoaspartic acid increases significantly with storage at 2–8
°C. Hydrophobic interaction chromatography was utilized to separate the isoaspartic variant in the intact state. Mass spectrometry including peptide mapping was employed to identify and confirm the exact location of the modification. Since this modification occurs in the complementarity determining region (CDR) it was found that binding is reduced. Therefore, three different analytical methods for regular analysis of this isomerization are evaluated. These methods include peptide mapping by LC–MS, HIC, and a protein isoaspartate methyltransferase assay. It was determined that HIC is the best method to regularly assay the level of isomerization in this monoclonal antibody.</description><subject>Analysis</subject><subject>Analytical, structural and metabolic biochemistry</subject><subject>Antibodies, Monoclonal - chemistry</subject><subject>Biological and medical sciences</subject><subject>Chromatography, Affinity - methods</subject><subject>Complementarity Determining Regions - chemistry</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>General pharmacology</subject><subject>Humans</subject><subject>Hydrophobic interaction chromatography</subject><subject>Isoaspartic Acid - chemistry</subject><subject>Isomerism</subject><subject>Isomerization</subject><subject>Mass Spectrometry - methods</subject><subject>Medical sciences</subject><subject>Monoclonal antibody</subject><subject>Peptide mapping</subject><subject>Peptide Mapping - methods</subject><subject>Pharmacology. Drug treatments</subject><subject>PIMT assay</subject><issn>1570-0232</issn><issn>1873-376X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2009</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkcuKFDEUhgtRnHH0EZRsxFW1SaUrqVqJDF4GBtwouAunkpPpNJWkTVJCP4MvbcoudCkcSOB85_b_TfOS0R2jTLw97o76kKKfdh2l424Nzh4112yQvOVSfH9c_72kLe14d9U8y_lIKZNU8qfNFRsHNkohrptfdwZDcdZpKC4GAsEQj5CXhL4mSLTE5Qj5BKk4TUA7Q2xM_kK7QMoBiY7-NP_hIblyJgYLJu-CCw8k4cNK1j5A7DLPZ3JYPATiY4h6jgHmOrO4KZrz8-aJhTnji-29ab59_PD19nN7_-XT3e37-1bvh7G0XBs2ajrtB953YKSYJloPHUVnwTLbUzn0PYV-QCaoNZ3tJGpJASnlOIDgN82bS99Tij8WzEV5lzXOMwSMS1aS76vEo1jJ_kLqFHNOaNUpOQ_prBhVqw3qqDYb1GqDWoOzWvdqm7BMHs2_qk33CrzeAMgaZpsgaJf_cl3HBBvHsXLvLhxWPX46TCprh0GjcQl1USa6_6zyG5p5rE8</recordid><startdate>20091115</startdate><enddate>20091115</enddate><creator>Dick, Lawrence W.</creator><creator>Qiu, Difei</creator><creator>Cheng, Kuang-Chuan</creator><general>Elsevier B.V</general><general>Elsevier</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>7X8</scope></search><sort><creationdate>20091115</creationdate><title>Identification and measurement of isoaspartic acid formation in the complementarity determining region of a fully human monoclonal antibody</title><author>Dick, Lawrence W. ; Qiu, Difei ; Cheng, Kuang-Chuan</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c489t-3cd19c0b48352ad76bb0570962faf1f5078550a58e160fd2f27ec70ae003e8a63</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2009</creationdate><topic>Analysis</topic><topic>Analytical, structural and metabolic biochemistry</topic><topic>Antibodies, Monoclonal - chemistry</topic><topic>Biological and medical sciences</topic><topic>Chromatography, Affinity - methods</topic><topic>Complementarity Determining Regions - chemistry</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>General pharmacology</topic><topic>Humans</topic><topic>Hydrophobic interaction chromatography</topic><topic>Isoaspartic Acid - chemistry</topic><topic>Isomerism</topic><topic>Isomerization</topic><topic>Mass Spectrometry - methods</topic><topic>Medical sciences</topic><topic>Monoclonal antibody</topic><topic>Peptide mapping</topic><topic>Peptide Mapping - methods</topic><topic>Pharmacology. Drug treatments</topic><topic>PIMT assay</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Dick, Lawrence W.</creatorcontrib><creatorcontrib>Qiu, Difei</creatorcontrib><creatorcontrib>Cheng, Kuang-Chuan</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>MEDLINE - Academic</collection><jtitle>Journal of chromatography. B, Analytical technologies in the biomedical and life sciences</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Dick, Lawrence W.</au><au>Qiu, Difei</au><au>Cheng, Kuang-Chuan</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Identification and measurement of isoaspartic acid formation in the complementarity determining region of a fully human monoclonal antibody</atitle><jtitle>Journal of chromatography. B, Analytical technologies in the biomedical and life sciences</jtitle><addtitle>J Chromatogr B Analyt Technol Biomed Life Sci</addtitle><date>2009-11-15</date><risdate>2009</risdate><volume>877</volume><issue>30</issue><spage>3841</spage><epage>3849</epage><pages>3841-3849</pages><issn>1570-0232</issn><eissn>1873-376X</eissn><abstract>Isomerization plays a key role in protein degradation. This isomerization is often difficult to detect by many protein characterization methods such as SDS-PAGE, SEC, and IEF. This work shows the identification of an isomerized aspartic acid residue in the CDR2 of the heavy chain of a fully human monoclonal antibody. This isoaspartic acid increases significantly with storage at 2–8
°C. Hydrophobic interaction chromatography was utilized to separate the isoaspartic variant in the intact state. Mass spectrometry including peptide mapping was employed to identify and confirm the exact location of the modification. Since this modification occurs in the complementarity determining region (CDR) it was found that binding is reduced. Therefore, three different analytical methods for regular analysis of this isomerization are evaluated. These methods include peptide mapping by LC–MS, HIC, and a protein isoaspartate methyltransferase assay. It was determined that HIC is the best method to regularly assay the level of isomerization in this monoclonal antibody.</abstract><cop>Amsterdam</cop><pub>Elsevier B.V</pub><pmid>19819766</pmid><doi>10.1016/j.jchromb.2009.09.031</doi><tpages>9</tpages></addata></record> |
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| ispartof | Journal of chromatography. B, Analytical technologies in the biomedical and life sciences, 2009-11, Vol.877 (30), p.3841-3849 |
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| source | MEDLINE; Access via ScienceDirect (Elsevier) |
| subjects | Analysis Analytical, structural and metabolic biochemistry Antibodies, Monoclonal - chemistry Biological and medical sciences Chromatography, Affinity - methods Complementarity Determining Regions - chemistry Fundamental and applied biological sciences. Psychology General pharmacology Humans Hydrophobic interaction chromatography Isoaspartic Acid - chemistry Isomerism Isomerization Mass Spectrometry - methods Medical sciences Monoclonal antibody Peptide mapping Peptide Mapping - methods Pharmacology. Drug treatments PIMT assay |
| title | Identification and measurement of isoaspartic acid formation in the complementarity determining region of a fully human monoclonal antibody |
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