蛋白质纯化制备工艺研究动力工程及工程热物理专业论文.docxVIP

北京化工大学硕士学位论文错的效果。 北京化工大学硕士学位论文 错的效果。 然后对Tf样品进行了表征及活性鉴定。通过凝胶过滤高效液相色 谱(HPSEC)检测Tf的结构完整性及是否存在聚集体；圆二色光谱鉴定结 果表明Tf的二级结构没有发生改变；通过质谱检测其分子量为78726； 全波长扫描结果发现在465 nin处有明显的特征吸收峰，表明Tf能结合 两个铁离子；抗菌活性测定结果表明Tf对大肠杆菌的生长有明显的抑 制作用。 最后在最优层析条件下进行了Tf的纯化中试，每批次得N-r 1 g左 右的Tf样品。Tf的纯度都达到了99％以上，并且收率较实验阶段有所 提高达到了80％以上。 本文建立了新型的Tf两步层析纯化工艺，该工艺收率好、纯度高、 适合工业放大，同时将废弃组分再利用节约了宝贵的血浆资源；为Tf 的功能性研究提供了便利。 关键词：Cohn组分IV，转铁蛋白，纯化，离子交换层析，疏水层析 万方数据 ABSTRACTSTUDY ABSTRACT STUDY ON PREPARATIoN AND PUIUFICATIoN OF PRoTEIN ABSTRACT The key technology of proteomics is protein purification．Transferrin(Tf) is one of the trace protein，the concentration of which is 2．5 g／L in human plasma．Tf has many important functions such as antibacterial function， transporting iron and it could also be the growth factor of cell multiplication, More important，Tf contributes great research value in the field of cancer treatment．The current thesis takes Tf as target protein，created a two—step chromatography method of purifying Tf from Cohn fraction IV,which was discarded in the cold ethanol precipitation process．Tf production experiences the characterization and activity evaluation and Tf scale—up experiments were under optimum conditions． In the first place，the components，operation and working process of AKTA explorer 1 00 protein purification system were introduced，which laid the foundation for the subsequent protein purification process． Secondly,two-stepchromatographymethod of pufffying Tf from Cohn fraction IV was researched．It was found that Yris—HCl buffer is not good for the centrifugal of Cohn fraction IV in pretreatment process，which III 万方数据 万方数据 万方数据 北京化工大学硕士学位论文influenced 北京化工大学硕士学位论文 influenced the later filtration，Thus，we decided to dissolve Cohn fraction IV in ultra pure water．According to the differences in isoelectric point and hydrophobicity,we choose the purification scheme of the combination of ion exchange chromatography and hydrophobic chromatography．After comparing two important factors，the chromatographic medi