Kanumuri Radha Madhavi, Bajji Chitra, Tummuru Rajesh R, Tatireddigari Venkat R R Arva, Mangamoori Lakshmi Narasu, Panati Kalpana, Narala Venkata Ramireddy
Virchow Research Centre, Hyderabad 500 055, India; Centre for Biotechnology, Jawaharlal Nehru Technological University, Kukatpally, Hyderabad 500 085, India.
Virchow Research Centre, Hyderabad 500 055, India.
Protein Expr Purif. 2015 Jul;111:61-7. doi: 10.1016/j.pep.2015.03.011. Epub 2015 Mar 27.
Recombinant human B-type natriuretic peptide (rhBNP) is a 32-amino acid peptide used to treat congestive heart failure. In this paper, we report a method for the increased production of rhBNP in Escherichia coli with high purity. hBNP was cloned with a short growth hormone fusion partner coupled with a unique acid-labile dipeptide linker to cleave the fusion protein to release the rhBNP. The recombinant fusion protein was expressed as an inclusion body (IB) and the fermentation process was optimized to produce on large scale. The IBs were recovered by cell lysis, and the pure IBs were directly treated with diluted acid to get the target peptide from the fusion protein and the resultant peptide was purified by reversed phase chromatography. The final purity of the rhBNP was more than 99% with yield of 50mg per liter of culture, which is ten times higher than the previous reports. The purified rhBNP exhibited specific biological activity similar to the standard peptide in producing cyclic-guanosine monophosphate.
重组人B型利钠肽（rhBNP）是一种用于治疗充血性心力衰竭的32个氨基酸的肽。在本文中，我们报告了一种在大肠杆菌中高效生产高纯度rhBNP的方法。hBNP与一个短的生长激素融合伴侣一起克隆，并带有一个独特的酸不稳定二肽接头，用于切割融合蛋白以释放rhBNP。重组融合蛋白以包涵体（IB）形式表达，并对发酵过程进行了优化以进行大规模生产。通过细胞裂解回收包涵体，将纯包涵体直接用稀酸处理以从融合蛋白中获得目标肽，所得肽通过反相色谱法纯化。rhBNP的最终纯度超过99%，每升培养物的产量为50mg，比以前的报道高出十倍。纯化的rhBNP在产生环磷酸鸟苷方面表现出与标准肽相似的特定生物活性。
