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Detection and Identification of the Vibrational Markers for the Quantification of Methionine Oxidation in Therapeutic Proteins.
Balakrishnan, Gurusamy; Barnett, Gregory V; Kar, Sambit R; Das, Tapan K.
Afiliación
  • Balakrishnan G; Molecular and Analytical Development , Bristol-Myers Squibb , 311 Pennington Rocky Hill Road , Pennington , New Jersey 08534 , United States.
  • Barnett GV; Molecular and Analytical Development , Bristol-Myers Squibb , 311 Pennington Rocky Hill Road , Pennington , New Jersey 08534 , United States.
  • Kar SR; Molecular and Analytical Development , Bristol-Myers Squibb , 311 Pennington Rocky Hill Road , Pennington , New Jersey 08534 , United States.
  • Das TK; Molecular and Analytical Development , Bristol-Myers Squibb , 311 Pennington Rocky Hill Road , Pennington , New Jersey 08534 , United States.
Anal Chem ; 90(11): 6959-6966, 2018 06 05.
Article en En | MEDLINE | ID: mdl-29741878
Methionine oxidation is a major degradation pathway in therapeutic proteins which can impact the structure and function of proteins as well as risk to drug product quality. Detecting Met oxidation in proteins by peptide mapping followed by liquid chromatography with mass spectrometry (LC-MS) is the industry standard but is also labor intensive and susceptible to artifacts. In this work, vibrational difference spectroscopy in combination with 18O isotopic shift enabled us to demonstrate the application of Raman and FTIR techniques for the detection and quantification of Met oxidation in various therapeutic proteins, including mAbs, fusion proteins, and antibody drug conjugate. Vibrational markers of Met oxidation products, such as sulfoxide and sulfone, corresponding to S═O and C-S═O stretching frequencies were unequivocally identified based 18O isotoptic shifts. The intensity of the isolated νC-S Raman band at 702 cm-1 was successfully applied to quantify the average Met oxidation level in multiple proteins. These results are further corroborated by oxidation levels measured by tryptic peptide mapping, and thus the confirmed Met oxidation levels derived from Raman and mass spectrometry are indeed consistent with each other. Thus, we demonstrate the broader application of vibrational spectroscopy to detect the subtle spectral changes associated with various chemical or physical degradation of proteins, including Met oxidation as well as higher order structural changes.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Sulfonas / Sulfóxidos / Proteínas Recombinantes de Fusión / Metionina / Anticuerpos Monoclonales Tipo de estudio: Diagnostic_studies Idioma: En Revista: Anal Chem Año: 2018 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Sulfonas / Sulfóxidos / Proteínas Recombinantes de Fusión / Metionina / Anticuerpos Monoclonales Tipo de estudio: Diagnostic_studies Idioma: En Revista: Anal Chem Año: 2018 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Estados Unidos