Vemula Sandeep, Vemula Sushma, Dedaniya Akshay, Kante Rajesh Kumar, Ronda Srinivasa Reddy
a Centre for Bioprocess Technology, Department of Biotechnology , K. L. E. F. University , Guntur , Andhra Pradesh , India.
b Department of Pharmacology, Kakatiya University , Warangal , Andhra Pradesh , India.
Prep Biochem Biotechnol. 2016 Aug 17;46(6):586-95. doi: 10.1080/10826068.2015.1084933.
The Fourier-transform infrared (FT-IR) spectroscopic approach has been employed to understand the recombinant human G-CSF (rhG-CSF) protein accumulation, secondary structure, and thermal stability in Escherichia coli grown under a temperature shift strategy (37 and 28°C) in various media formulations. The choline + sodium pyruvate (37°C) and sodium pyruvate (28°C) formulations have shown the highest inclusion body (IB) accumulation of 0.41 and 0.46 mg/mL, respectively. Furthermore, insights on the structure of the rhG-CSF within IBs and intact cells have been investigated through secondary structure analysis. Thermal stability experiments were also carried out to explain the pattern of the second derivative structure of rhG-CSF. The studies showed that choline + sodium pyruvate formulation has preserved the protein secondary structure even at 82°C. Overall, the FT-IR spectroscopic technique can also be adopted to accelerate the characterization of other recombinant therapeutic proteins of E. coli origin.
傅里叶变换红外（FT-IR）光谱方法已被用于了解在不同培养基配方下采用温度转换策略（37和28°C）培养的大肠杆菌中重组人粒细胞集落刺激因子（rhG-CSF）蛋白的积累、二级结构和热稳定性。胆碱 + 丙酮酸钠（37°C）和丙酮酸钠（28°C）配方分别显示出最高的包涵体（IB）积累量，为0.41和0.46 mg/mL。此外，通过二级结构分析研究了包涵体和完整细胞内rhG-CSF的结构。还进行了热稳定性实验以解释rhG-CSF二阶导数结构的模式。研究表明，胆碱 + 丙酮酸钠配方即使在82°C时也能保留蛋白质二级结构。总体而言，FT-IR光谱技术也可用于加速对其他大肠杆菌来源的重组治疗性蛋白质的表征。
