Characterization of the primary structure of TNK-tissue plasminogen activator using LC-MS.
- Author:
Lei TAO
1
;
You-Xue DING
;
Ying GUO
;
Chun-Ming RAO
;
Jun-Zhi WANG
Author Information
1. Key Laboratory of the Ministry of Health for Research on Quality and Standardization of Biotech Products, National Institute for Food and Drug Control, Beijing 100050, China.
- Publication Type:Journal Article
- MeSH:
Amino Acid Sequence;
Chromatography, Liquid;
Glycosylation;
Mass Spectrometry;
Molecular Weight;
Protein Processing, Post-Translational;
Tissue Plasminogen Activator;
chemistry
- From:
Acta Pharmaceutica Sinica
2013;48(6):896-900
- CountryChina
- Language:Chinese
-
Abstract:
The primary structure of TNK-tissue plasminogen activator (TNK-tPA) was characterized using liquid chromatography-mass spectrometry (LC-MS). Firstly, the molecular mass of deglycosylated protein was measured. Then peptide mass mapping and MS/MS of the reduced, alkylated and trypsin-digested sample were tested and analyzed so as to verify its amino acid sequence and identify post-translational modifications. Results show that the amino acid sequence was consistent with designed structure; about 5% of M207 was oxidized; T61 was fucosylated with -80% occupancy; N103, N448 and N184 (-15% occupancy) were glycosylated with complex-type oligosaccharides. LC-MS coupled with proper sample pretreatment is approved to be a rapid and powerful approach to characterize the primary structure of TNK-tPA.
