驴血清α1酸性糖蛋白生物信息学分析与纯化鉴定研究

doi:10.16431/j.cnki.1671-7236.2021.08.003

摘要/Abstract

摘要： 研究旨在对驴α1酸性糖蛋白（alpha-1-acid glycoprotein，α1-AGP）进行生物信息学预测和分析，并建立一种高效提纯驴α1-AGP的方法。通过生物信息学在线软件分析驴α1-AGP的基本理化性质，并对其二、三级结构进行预测分析；用阳离子层析-凝胶过滤层析联用的方法对其进行纯化，经SDS-PAGE后用基质辅助激光解吸电离飞行时间质谱（matrix-assisted laser desorption ionization-time offlight mass spectrometry，MALDI-TOF-MS）进行鉴定。结果显示，驴α1-AGP由201个氨基酸组成，理论等电点（pI）为4.99，有5个潜在N-糖基化位点，亲水性总平均值（GRAVY）为-0.386；二级结构中α-螺旋和β-折叠有序出现，占比88.56%；利用GMQE值和QMEAN的z值验证三级结构预测所得模型在合理范围之内。阳离子交换层析-凝胶过滤层析纯化后得到50 mL蛋白液，SDS-PAGE后得到清晰蛋白条带，经MALDI-TOF-MS鉴定后于NCBI数据库检索，驴α1-AGP得分最高且远高于其他蛋白，证明驴α1-AGP为纯化后蛋白液中主要组分。本研究提出一种新的纯化鉴定驴α1-AGP方法，具有纯化率高、操作简便、所需样品少等优点，为驴α1-AGP的后续生物学研究和应用奠定了理论基础。

关键词: 驴; α1酸性糖蛋白(α1-AGP); 生物信息学; 纯化

Abstract: This study was aimed to carry out the bioinformatics prediction and analysis of alpha-1-acid glycoprotein(α1-AGP) of donkey, and an efficient purification method of α1-AGP from donkey was established.Analysis of donkey by bioinformatics online software.The basic physical, chemical properties and the secondary and tertiary structures of α1-AGP were predicted.According to the results of the analysis, the method of cation chromatography gel filtration chromatography was used to purify donkey α1-AGP, after SDS-PAGE, matrix assisted laser desorption ionization-time offlight mass spectrometry (MALDI-TOF-MS) was used to identify donkey α1-AGP.The results showed that donkey α1-AGP was stable and hydrophilic, composed of 201 amino acids, the theoretical isoelectric point(pI) was 4.99, there were 5 potential N-glycosylation sites, and the average value of hydrophilicity(GRAVY) was -0.386.α-helix and β-sheet appeared orderly in the secondary structure, accounting for 88.56%, the GMQE value and the Z value of QMEAN were used to verify that the model was in a reasonable range.After purification by cation exchange chromatography and gel filtration chromatography, a total of 50 mL protein solution was obtained, and a clear protein strip was obtained after SDS-PAGE.After MALDI-TOF-MS and retrieved from NCBI database, the protein with the highest score was donkey α1-AGP, which was much higher than other proteins, indicating that α1-AGP was the main component in the purified protein.In this study, we proposed a new method for the purification and identification of α1-AGP from donkeys, which had the advantages of high purification rate, simple operation and less samples needed, and laid a theoretical foundation for the subsequent biological research and application of α1-AGP from donkeys.

Key words: donkey; alpha-1-acid glycoprotein (α1-AGP); bioinformatics; purification

中图分类号:

S822

费雅麒, 汤驰, 邓亮. 驴血清α1酸性糖蛋白生物信息学分析与纯化鉴定研究[J]. 中国畜牧兽医, 2021, 48(8): 2705-2712.

FEI Yaqi, TANG Chi, DENG Liang. Bioinformatics Analysis,Purification and Identification of Alpha-1-acid Glycoprotein in Serum of Donkey[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(8): 2705-2712.

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