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目的 :研究重组人肝细胞生成素 (hepatopoietin ,HPO)的理化性质。方法 :在大肠杆菌中表达的人HPO ,经凝胶过滤和离子交换柱层析进行纯化 ;通过还原型SDS 聚丙烯酰胺凝胶电泳(SDS PAGE)、非还原型SDS PAGE及基质辅助激光解析飞行时间质谱 (MAIDI TOF MS)等方法分别测定了人HPO的相对分子质量 ;采用等电聚焦电泳测定了人HPO的等电点 ;采用内源荧光光谱和圆二色光谱法研究了人HPO的稳定性。结果与结论 :复性后的重组人HPO主要以双体形式存在 ,其相对分子质量为 30 0 0 0 ,等电点在 5.2～ 6 .0之间。在 70℃以下、pH 2 .6以上 ,人HPO分子内部构象基本不变
Objective: To study the physicochemical properties of recombinant human hepatopoietin (HPO). Methods: Human HPO expressed in Escherichia coli was purified by gel filtration and ion exchange column chromatography. The recombinant HPO was purified by SDS PAGE, non-reducing SDS PAGE and matrix-assisted laser desorption Time mass spectrometry (MAIDI TOF MS) were used to determine the relative molecular mass of human HPO respectively. The isoelectric point of human HPO was determined by isoelectric focusing electrophoresis. The stability of human HPO was studied by endogenous fluorescence spectroscopy and circular dichroism spectroscopy Sex. RESULTS AND CONCLUSION: The renatured recombinant human HPO mainly exists as a double body with a relative molecular mass of 30 000 and an isoelectric point of 5.2-6.0. Below 70 ° C, above pH 2.6, the internal conformation of the human HPO molecule remains essentially unchanged