title: [Characterization of the primary structure of TNK-tissue plasminogen activator using LC-MS].
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.
authors.affiliation: 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.
authors.forename: Lei
authors.initials: L
authors.lastname: Tao
authors.forename: You-Xue
authors.initials: YX
authors.lastname: Ding
authors.forename: Ying
authors.initials: Y
authors.lastname: Guo
authors.forename: Chun-Ming
authors.initials: CM
authors.lastname: Rao
authors.forename: Jun-Zhi
authors.initials: JZ
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D002853:Chromatography, Liquid; D006031:Glycosylation; D013058:Mass Spectrometry; D008970:Molecular Weight; D011499:Protein Processing, Post-Translational; D010959:Tissue Plasminogen Activator / Q000737:chemistry*
chemical_list: C104096:TNK-tissue plasminogen activator; D010959:Tissue Plasminogen Activator