Establishment of a high-throughput analysis method for the identification of key proteases of peptide drugs in vitro
10.11665/j.issn.1000-5048.20190312
- VernacularTitle:多肽药物关键水解酶体外高通量分析方法的建立
- Author:
Fan ZHANG
1
;
Hong TIAN
;
Wenbing YAO
Author Information
1. 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
- Publication Type:Journal Article
- Keywords:
peptide;
enzymatic catabolism;
HPLC;
proteases;
high-throughput analysis;
pramlintide
- From:
Journal of China Pharmaceutical University
2019;50(3):352-356
- CountryChina
- Language:Chinese
-
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.
