液质联用对重组胰高血糖素样肽-1-Fc融合蛋白中的制品相关蛋白进行定性定量分析

目的：对重组胰高血糖素样肽-1-Fc融合蛋白（rGLP-1-Fc）的制品相关蛋白进行定性及定量分析。方法：利用液质联用法，对rGLP-1-Fc原液中制品相关蛋白的修饰类型进行鉴定；通过胰蛋白酶酶切液质肽图的方法，对修饰位点进行鉴定；采用反相超高效液相色谱法，测定3批原液中制品相关蛋白的比例。结果：该蛋白原液中主要存在3种制品相关蛋白；液质联用检测结果显示：1种为氧化及糖化的修饰产物，另外2种为一条二硫键解链导致的变异体；液质肽图分析发现氧化及糖化发生在第28位赖氨酸残基；对3批原液进行了检测，3种制品相关蛋白的平均含量分别为（2.37±0.04）%、（9.07±0.20）%和（4.03±0.07）%。结论：利用液质联用技术对rGLP-1-Fc原液中的制品相关蛋白进行了初步的定性、定量分析，测定结果可为同类产品的制品相关蛋白分析提供技术支持。

Objective: To identify and quantify the product-related proteins in the bulk of recombinant GLP-1-Fc fusion protein (rGLP-1-Fc). Methods: Modifications of the product-related proteins were identified using ultra performance liquid chromatography-mass spectrometry (UPLC-MS). The modified site was determined by trypsin-digested peptide LC-MS mapping. The percentages of product-related proteins in three batches of bulk were detected by reversed-phase UPLC. Results: Three types of product-related proteins were found in the bulk,and results of UPLC-MS showed that one was oxidized and glycated protein and the other two were the variants caused by breaking of one disulfide bond. The modification site of oxidation and glycation was K28 determined by peptide mass mapping. The average percentage contents of the three product-related proteins in three batches of bulk were (2.37±0.04)%,(9.07±0.20)% and (4.03±0.07)%,respectively. Conclusion: The product-related proteins in rGLP-1-Fc were identified and quantified preliminarily by UPLC-MS,which could provide technical support for product-related protein analysis of similar products.

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