重组血红蛋白的原核表达、纯化和鉴定  

Prokaryotic expression,purification and identification of recombinant hemoglobin

作　　者：杨海波[1] 廖春丽[1] 朱涛[1] 王莲哲[1] 陈兰英[1] 

机构地区：[1]河南城建学院生物工程系,平顶山467044

出　　处：《重庆医学》2010年第21期2889-2891,共3页Chongqing medicine

基　　金：国家自然科学基金资助项目(30470877);河南省重点科技攻关项目(092102310006)

摘　　要：目的利用大肠杆菌表达载体,表达血红蛋白(Hb)基因,并进行纯化及鉴定。方法将原核表达载体pHE 2-Hb转化入E.Coli JM109,经IPTG诱导表达;应用阴离子交换层析、阳离子交换层析分级纯化目的蛋白,用Western blot鉴定纯化出的目的蛋白。结果 SDS-PAGE分析表明,表达产物的相对分子质量约为67 kD,与理论值相一致;应用阴离子交换层析、阳离子交换层析分级纯化目的蛋白,Western blot检测表明得到纯度较高的目的蛋白。结论利用大肠杆菌表达系统,获得了较高纯度的目的蛋白,为进一步研究Hb的生物传氧机制研究和以Hb为基础载氧药物的研究提供重要基础和研究依据,并将为血液替代品和相关疾病的治疗研究奠定研究基础。Objective To construct a prokaryotic expression vector for the expression of hemoglobin to purify and identity.Methods Prokaryotic expression vector pHE 2-Hb was expressed in E.Coli JM109.The expressed protein was identified by SDS-PAGE analysis.The target protein was cation-exchange chromatography and anion-exchange chromatography.Results The SDS-PAGE showed that the relative molecular mass of this protein was consistent with the predicted value.The protein was expressed in E.Coli JM109 after the prokaryotic expression vector pHE 2-Hb successfully constructed,and the size of the protein was 67 kD,which was consistent with the theoretical data.Conclusion The successfully constructed prokaryotic expression vector pHE 2-Hb,and the purified Hb protein can not only provide an important foundation to the study of more about the oxygen transfer mechanism of Hb,and Hb-based oxygen-carrying drugs,but also provide research proofs to the blood substitutes and the treatment of related diseases.

关 键 词：血红蛋白 原核表达 纯化 

分 类 号：Q78[生物学—分子生物学]