电泳/质谱技术研究鲨鱼肝铁蛋白及亚基多聚体特性

改良鲨鱼肝铁蛋白(liver ferritin ofsphyrna zygaena,SZLF)分离技术,并结合非变性梯度聚丙烯酰胺凝胶电泳(NGPAGE)制备质谱纯SZLF,以维系铁蛋白分子和它的亚基之间的作用类型,供研究铁蛋白结构与功能。实验结果表明,SZLF由分子量约为20kDa的单类型亚基组成。SZLF和它的亚基均组成不同的聚合体。聚合体类型和聚合数目与铁蛋白亚基和分离介质有关。MALDI-TO F质谱仪的激光和基质协同能有效解吸SZLF中的亚基成为准分子离子,并供质量分析,其亚基特征质谱峰m/z值分别为10889.35和22030.45,确定为带双电荷M 2H]2 和单电荷M H] 的SZLF亚基分子量。SDS-PAGE和变性IEF技术研究指出,形成不同聚合态的SZLF,其分子之间的相互作用强度高于SZLF亚基自身,难以通过MALDI-TOF质谱技术测定其亚基的结构信息。尽管SZLF由单类型亚基组成,但它能通过聚合体和自身亚基之间的相互作用差异性发挥极其重要的生理功能。

Polymer Characteristics of Shark Liver Ferritin and Its Subunits by Electrophoresis and Mass Spectrometry

An analytical technology from the Liver ferritin of Sphyrna zygaena(SZLF)has been modified for maintaining original interaction types both ferritin and its subunits for studying on the structure and function.A non-denaturing degrade polyacrylamide gel electrophoresis(ND-PAGE)was used to produce SZLF with high purity for analysis of mass spectrometry(MS).It was found that there were different polymers both SZLF and its subunits.The types and numbers of polymers were connected with the protein subunits and the separation medium.The laser come from the MALDI-TOF MS and matrix has capacities for making the subunits of ferritin into molecular ions for mass analysis,indicating that m/s of two representive peaks are 10889.35 and 22030.45,called M 2H]2 with double charge and M H] with single charge.Both SDS-PAGE and NDPAGE revealed that the interaction intensity of SZLF with different polymer are higher than that of single protein itself,which resulted in the structural information of subunit from the SZLF were difficult to be determined by MALDI-TOF MS.Even the protein consists of single subunit type,the polymer interaction both SZLF and its subunits still play an important role in carrying out its physiological functions.