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多肽药物关键水解酶体外高通量分析方法的建立

张凡, 田浤, 姚文兵

张凡, 田浤, 姚文兵. 多肽药物关键水解酶体外高通量分析方法的建立[J]. 中国药科大学学报, 2019, 50(3): 352-356. DOI: 10.11665/j.issn.1000-5048.20190312
引用本文: 张凡, 田浤, 姚文兵. 多肽药物关键水解酶体外高通量分析方法的建立[J]. 中国药科大学学报, 2019, 50(3): 352-356. DOI: 10.11665/j.issn.1000-5048.20190312
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ZHANG Fan, TIAN Hong, YAO Wenbing. Establishment of a high-throughput analysis method for the identification of key proteases of peptide drugs in vitro[J]. Journal of China Pharmaceutical University, 2019, 50(3): 352-356. DOI: 10.11665/j.issn.1000-5048.20190312
Citation: ZHANG Fan, TIAN Hong, YAO Wenbing. Establishment of a high-throughput analysis method for the identification of key proteases of peptide drugs in vitro[J]. Journal of China Pharmaceutical University, 2019, 50(3): 352-356. DOI: 10.11665/j.issn.1000-5048.20190312
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多肽药物关键水解酶体外高通量分析方法的建立

  • 中国药科大学生命科学与技术学院 江苏省生物药物成药性研究重点实验室

基金项目: 国家自然科学基金资助项目(No.81430082)

Establishment of a high-throughput analysis method for the identification of key proteases of peptide drugs in vitro

摘要

    摘要
  • 摘要: 为了建立测定多肽药物体内代谢关键水解酶的高通量分析方法，本研究基于高效液相色谱技术，确定通用的体外酶解反应条件为:pH 7.8或9.0，缓冲体系为0.01 mol/L PBS或50 mmol/L Tris缓冲液，实现对多肽药物关键水解酶的统一高通量体外检测。利用该方法对多肽药物普兰林肽的关键水解酶进行分析，研究结果显示，基肽释放相关酶5和二肽基肽酶4对普兰林肽的水解作用最强，与微量热泳动分析结果一致。因此，本研究所建立的方法可用于分析多肽药物的关键水解酶，为优化多肽药物的酶稳定性提供方法参考与指导。
    Abstract: In order to establish an effective analytical method to detect the key proteases which affect the metabolism and the plasma half-life of peptides in vivo, a method to analyze the key proteases of peptide drugs based on high performance liquid chromatography in vitro was established. The general enzymatic reaction conditions in vitro were as follows: pH was 7. 8 or 9. 0 and the buffer system was 0. 01 mol/L PBS or 50 mmol/L Tris buffer. The results of pramlintide detected by this method showed that kallikrein-related peptidase 5 and dipeptidyl peptidase 4 had the strongest hydrolysis on pramlintide. The result was consistent with that determined by microscale thermophoresis, which indicated that kallikrein-related peptidase 5 and dipeptidyl peptidase 4 were the key proteases of pramlintide. This analytical method provides the basis for high-throughput stability screening of peptides and can be used to analyze key proteases of peptide drugs. It can also provide guidance for optimizing the protease stability of peptide drugs.
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    • 参考文献(14)
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  • 刊出日期:  2019-06-24

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