Structure-based antigenic epitope and PEGylation improve the efficacy of staphylokinase

Microbial Cell Factories

Lei Bai¹  Jianzhong Zhou¹  Wei Yang²  Qiangsi Niu³  Deqiang Wang⁴  Xin Jin⁴  Yueyuan Shi⁴  Ailong Huang⁴  Shilei Wang⁴  Yanying Xu⁴ 

[1] Department of Cardiology, The First Affiliated Hospital of Chongqing Medical University;Department of Laboratory Medicine, Hospital of Zhejiang;Department of Laboratory Medicine, The First Affiliated Hospital of Chongqing Medical University;Key Laboratory of Molecular Biology for Infectious Diseases (Ministry of Education), Institute for Viral Hepatitis, Department of Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University;

关键词: Staphylokinase;    PEGylation;    Immunogenicity;   

DOI  :  10.1186/s12934-017-0801-y

来源: DOAJ

【 摘 要 】

Abstract Staphylokinase (Sak) holds promise for use in thrombolytic therapy for acute myocardial infarction. However, its immunogenicity is a major disadvantage under clinical conditions. PEGylation has become a sophisticated method to decrease that immunogenicity. In this report, according predicted epitope from the active center, five residues, including Gly79, Leu82, Lys84, Ala97, and Arg104 have been mutant as cysteine for mono PEGylation, respectively. According to the relative immunogenicity of Sak or its PEGylation derivatives, the amount of specific anti-Sak IgG antibodies elicited by PEGylation proteins, including C79G, C82L, C84K, C97A, and C104R in BALB/c mice decreased by approximately 15–75% each. PEGylated Sak derivatives showed a decrease of up to 75% in the immune reactivity in PEG-Sak-C104R. Thrombelastography experiments showed that two PEG-conjugated derivatives, PEG-Sak-C97A (Ly30, 68.14 ± 2.51%) and PEG-Sak-C104R (Ly30, 66.49 ± 5.97%), the LY30 of PEG-Sak-C97A, and PEG-Sak-C104R produced values very similar to those of wild-type Sak. The fibrin plate assays showed the bioactivity of PEG-Sak-C104R to exhibit the most activity approximately as much as urokinase (diameter of halo pattern, 18.6 ± 1.06 mm) and tPA (diameter of halo pattern, 17.2 ± 0.49 mm). The Sak PEGylation derivative PEG-Sak-C104R was also selected for further in vivo activity experimentation. The thrombolytic ability of PEG-Sak-C104R is a little lower than wild-type Sak, whereas, this PEGylated protein retained high activity suitable for thrombolytic therapy. Collectively, with the in vivo and in vitro experiments, the present study suggests that site mutant PEGylation, PEG-Sak-C104R, is a suitable type of PEGylation for clinical applications. Further optimization would help maintain the bioactivity and decrease the immunogenicity of staphylokinase.

【 授权许可】

Unknown