Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • harvard1
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Conformational studies of a monoclonal antibody, IgG1, by chemical oxidation: Structural analysis by ultrahigh-pressure LC-electrospray ionization time-of-flight MS and multivariate data analysis
Department of Analytical Chemistry, Stockholm University.
Show others and affiliations
2008 (English)In: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 380, no 2, 155-163 p.Article in journal (Refereed) Published
Abstract [en]

We describe the development of a method in which protein oxidation by H2O2 followed by ultrahigh-pressure liquid chromatography (UHPLC) coupled with electrospray ionization time-of-flight mass spectrometry (ESI-ToFMS) and multivariate analysis are used to detect alterations in conformational states of proteins. In the study reported here, an IgG1 monoclonal antibody in native and denatured conformational states was oxidized by treatment with hydrogen peroxide. Peptide fragments generated by tryptic digestion were then analyzed by UHPLC-ESI-ToFMS. After reducing noise and extracting peaks from the LC-MS data using MzExplorer, software developed in-house and based on Matlab, we were able to distinguish peptides arising from the native and denatured states of the oxidized protein by principal component analysis. Peptides containing residues, which are inclined to undergo oxidation, such as methionine, are founded to be particularly important in this approach. We believe that the methodology could facilitate attempts to characterize the conformational states of recombinant monoclonal antibodies and other proteins.

Place, publisher, year, edition, pages
2008. Vol. 380, no 2, 155-163 p.
Keyword [en]
Mass spectrometry, Methionine, Peptide mapping, Principal component analysis, Protein conformation, Recombinant monoclonal antibody, Ultrahigh-pressure liquid chromatography
National Category
Analytical Chemistry
Identifiers
URN: urn:nbn:se:kth:diva-87062DOI: 10.1016/j.ab.2008.05.054ISI: 000258313100001PubMedID: 18577369OAI: oai:DiVA.org:kth-87062DiVA: diva2:501351
Note
QC 20120214Available from: 2012-02-14 Created: 2012-02-14 Last updated: 2017-12-07Bibliographically approved
In thesis
1. Methods for structural studies of an antibody, screening metabolites in rat urine and analysis of spent cell cultivation media using LC/ESI-MS and chemometrics
Open this publication in new window or tab >>Methods for structural studies of an antibody, screening metabolites in rat urine and analysis of spent cell cultivation media using LC/ESI-MS and chemometrics
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis describes bioanalytical methods for generating fingerprints of biological systems for extracting relevant information with (protein) drugs in focus. Similarities and differences between samples can reveal the hidden relevant information, which can be used to optimize the production and facilitate the quality control of such protein drugs during their development and manufacture. Metabolic fingerprinting and multivariate data analysis (MVDA) can also facilitate early diagnosis of diseases and the effects and toxicity of drugs.

Currently, several protein drugs are available on the global market. Nevertheless, despite, the success of such biotherapeutics significant challenges remain to be overcome in maintaining their stability and efficacity throughout their production cycle and long-term storage. The native structure and functional activity of therapeutic proteins is affected by many variables from production to delivery, incl. variables assoc. with conditions in bioreactors, purification, storage and delivery. Thus, part of the work underlying this thesis focused on structural analysis of a protein drug using chemical labeling, peptide mapping, and evaluation of the charge state distributions of the whole protein generated by ESI. The other part focuses on non-targeted metabolomics with a view to optimizing the cell cultivation process and assessment of the drug’s toxicity. A combination of appropriate analytical methods and MVDA is needed to find markers that can facilitate optimization of the cultivation system and expression of the target proteins in early stages of process development. Rapid methods for characterizing the protein drugs in different stages of the process are also required for quality control.

In order to obtain high quality fingerprints analytical separation techniques with high resolution (such as HPLC or UHPLC) and sensitive analytical detection techniques (such as ESI, quadrupole or TOF MS) have been used, singly or in combination.

Place, publisher, year, edition, pages
Stockholm University, 2009. 74 p.
Keyword
Screening, Fingerprinting, metabolomics, Protein drugs, Mammalian Cell lines, LC/ESI-MS, chemometrics
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:kth:diva-87121 (URN)978-91-7155-897-8 (ISBN)
Note
QC 20120215Available from: 2012-02-15 Created: 2012-02-14 Last updated: 2012-02-15Bibliographically approved

Open Access in DiVA

No full text

Other links

Publisher's full textPubMed

Search in DiVA

By author/editor
Zamani, Leila
In the same journal
Analytical Biochemistry
Analytical Chemistry

Search outside of DiVA

GoogleGoogle Scholar

doi
pubmed
urn-nbn

Altmetric score

doi
pubmed
urn-nbn
Total: 64 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • harvard1
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf