CAJ | 학술논문

为了获得兼具“高弹性”和“硅沉积”性能的新型功能生物材料，将具有高弹性的Resilin蛋白与具有硅沉积作用的R5肽进行了融合，研究其在大肠杆菌中的表达，建立了简易的纯化方法。首先，将果蝇弹性蛋白基因CG15920序列与硅藻R5肽基因序列进行拼接，经过稀有密码子优化后合成重组基因resilin-r5。然后将其插入原核表达载体pET28a，转化大肠杆菌BL21（DE3）pLysS获得表达菌株，并通过IPTG进行诱导表达。最后以盐析加热法和Ni离子柱亲和层析法纯化带组氨酸标签的重组融合蛋白。最终实现Resilin-R5融合蛋白的高效表达，产量达到120 mg· L-1发酵液，为“高弹性-硅沉积”融合蛋白Resilin-R5的性能表征和创新应用奠定了基础。
위료획득겸구“고탄성”화“규침적”성능적신형공능생물재료，장구유고탄성적Resilin단백여구유규침적작용적R5태진행료융합，연구기재대장간균중적표체，건립료간역적순화방법。수선，장과승탄성단백기인CG15920서렬여규조R5태기인서렬진행병접，경과희유밀마자우화후합성중조기인resilin-r5。연후장기삽입원핵표체재체pET28a，전화대장간균BL21（DE3）pLysS획득표체균주，병통과IPTG진행유도표체。최후이염석가열법화Ni리자주친화층석법순화대조안산표첨적중조융합단백。최종실현Resilin-R5융합단백적고효표체，산량체도120 mg· L-1발효액，위“고탄성-규침적”융합단백Resilin-R5적성능표정화창신응용전정료기출。
A fusion protein Resilin-R5 which was expected to have a superiority of high elasticity and silicon deposi-tion was designed and expressed in Escherichia coli.At the same time, a facile purification method was built to ob-tain the new fusion protein.Firstly, the recombinant resilin-r5 gene, which constructed by fusing the diatoms R5 peptide gene sequences to the 3’ end of Drosophila melanogaster resilin gene sequences, was synthesized after a rare codon optimization.Then, the new gene was inserted into the prokaryotic expression vector pET 28a.The expression vector was transformed into E.coli BL21(DE3)pLysS strain, and the transformant was cultured in an auto-induced medium to get resilin-r5 gene expressed.At last, the His-tagged recombinant fusion protein was purified by‘salting-out and heating'method and Ni ion column affinity chromatography .The efficient expression of the fusion Resilin-R5 reached 120 mg per liter culture .A solid foundation was built for characterization and innovative applications of the fusion protein Resilin-R5 which has high elasticity and silicon deposition ability .