Comparison of largeª²scale preparation of recombinant human BMPª²4 and BMPª²2 mature peptide expressed in E.coli

    WANG Tao, CHEN Suª²Min ,CHEN Nanª²Chun, ZHAO Weiª²Qin£¬ZHANG Xiaoª²Nan

    Department of Biochemistry and Molecular Biology, School of Basic Medicine, Fourth Military Medical University, Xi¬ðan 710033, China

    ¡¾Abstract¡¿ AIM: To prepare pure human bone morphogenetic protein mature peptide (rhBMPª²4m and rhBMPª²2m)in large scale. METHODS: The rhBMPª²4m and rhBMPª²2m engineering strains were fermented and induced for expression in 15L NBS fermenter respectively and the bacterial cells were collected by centrifugation. Then the bacterial cells were lyzed and the resulting inclusion bodies were washed for 4 times. After obtaining data from preª²experiments, all inclusion bodies were dissolved in 8 mol/L urea buffer and loaded on SPª²Sepharose FF column. RESULTS: At the end of fermentation, A600 nm values of the two bacterial cultures were 28.8 and 26.3. SDSª²PAGE analysis showed that rhBMPª²4m occupied 40% of the total bacterial proteins while that of rhBMPª²2m was 47.2%. rhBMPª²4m and rhBMPª²2m accounted for 82.9% and 84.5% of the total proteins of the inclusion bodies respectively. After loaded on SPª²Sepharose FF column, rhBMPª²4m and rhBMPª²2m were eluted out in 0.35 and 0.20 mol/L NaCl fractions and the purity reached 96% and 95% with the harvest of 1.30 and 1.25 g/L fermentation liquid and the recovery rate were 34.33% and 34.81%, respectively. CONCLUSION: It could use the same fermentation program and the similar techniques to prepare pure rhBMPª²4m and rhBMPª²2m in large scale. ......