Purification And Characterization Of Rabbit Growth Hormone

As a therapeutic protein drug for growth hormone deficiency desease, humangrowth hormone has been used for many years. Meanwhile, the research regardingapplication of human growth hormone has never been stopped. It is very important toset up a proper animal model for the research. Rabbit is one of the ideal candidates.So we set up a process to express fusion rabbit growth hormone in E.Coli and purifyin this study. Mature rabbit growth hormone was obtained for the first time in theworld.Expressive vector pET11a and pET32a with rabbit growth hormone gene insertwere transformed into E.coli BL21(DE3) respectively. After IPTG induction thefusion protein MEAE-rabGH and Trx-D4K-rabGH were expressed in inclusionbodies. The expression level was 47mg/L·OD and 122 mg/L·OD. To obtain solubleexpression, low temperature culture condition and low concentration of IPTGinduction were tried. But no anticipated soluble expression was obtained. So thestudy had to be started from inclusion bodies.After inclusion bodies isolation, inclusion body solubilization, refoldingcondition and protease digestion condition were optimized and optimal conditionwas obtained. Using the optimal process, mature rabGH was recoverd at a purity ofmore than 95% with yield of 5.8%(Trx-D4K-rabGH) and 8.9%(MEAE-rabGH)respectively.The purified rabGH was determined intact molecule weight by MALDI-TOFMS.After trypsin digestion, a mass spectrum piptides map was obtained and thedisulphide bonds were Cys52-Cys163 and Cys180-Cys188. Cell based assay showed thatthe purified rabGH had biological activity.