HPLC用于重组L-天门冬酰胺酶Ⅱ的纯度分析与肽图谱测定

Purity Check and Tryptic Peptide Mapping of RecombinantL-Asparaginase by HPLC

目的：为了对重组Ｌ－天门冬酰胺酶Ⅱ进行肽图谱测定，以比较不同来源产品间一级结构的一致性。方法：运用梯度洗脱／反相高效液相色谱法对样品进行纯度检查和肽图谱测定，并提出了简单的三氯乙酸变性方法，以解除蛋白质抗胰蛋白酶水解的能力。结果：建立了重组Ｌ－天门冬酰胺酶Ⅱ的三氯乙酸变性／胰蛋白酶水解测定肽图谱的ＨＰＬＣ方法，肽图谱的比较反映出不同样品间的一级结构间存在差异。结论：建立的肽图谱测定方法，过程简便，便于常规测定，样品间的结构差异值得进一步研究。

Objective: To check the purity and characterize the peptide map of recombinant L-asparaginase expressed in Escherichia coli,so as to validate the primary structural identity of several samples from various sources. Method: Gradient elution/RP-HPLC was used.Prior to tryptic digest,a denaturing process with trichloroacetic acid was carried out to eliminate the protease-resistance from the protein. Results: Structural difference among several samples has been shown according to the peptide maps thus obtained. Conclusion: The method described was simple and easy-to-use.It is necessary to perform further research to exploit the primary strucutral difference of the protein.