Expression and production of recombinant scorpine as a potassium channel blocker protein in Escherichia coli.
Roghayyeh SeyfiValiollah BabaeipourMohammad Reza MofidFatemeh Abarghooi KahakiPublished in: Biotechnology and applied biochemistry (2018)
Scorpine is a cationic protein from the venom of Pandinus imperator, belonging to potassium channel blocker family, which has been shown to have antibacterial, antiviral, and antiplasmodia activities. In the present study, a pET-44a vector containing scorpine synthetic gene with T7 Promoter (pET 44a-His6-Nus-His6-tev-scorpine) was transferred into Escherichia coli Rosetta-gami B (DE3) for soluble expression of the protein in the cytoplasm and its overproduction. After confirming recombinant scorpine peptide expression using SDS-PAGE and Western blot, augmentation of production was performed during two stages. At first, effects of three parameters including carbon source concentration of medium, temperature, and induction time were investigated in terrific broth (TB) medium. Afterward, the overexpression was performed by response surface methodology in TB + glucose. Under the optimized conditions, the highest production of 3.5 g/L in the TB + glucose medium (7.5 g/L glucose, induction at OD600 = 3.5 and 25 °C) was increased to 4.1 g/L in TB medium (2.5 g/L glycerol, induction at OD600 = 0.7 and 25 °C). Then, in order to increase the amount of protein production, effects of carbon concentration in the fermenter under the primary optimized condition was investigated. The amount of produced recombinant protein increased from 0.12 to 2.1 g/L.H. The results were similar to previous studies on optimizing and increasing the production of recombinant protein and in particular recombinant scorpine.
Keyphrases
- escherichia coli
- binding protein
- poor prognosis
- protein protein
- amino acid
- computed tomography
- dna methylation
- cell free
- small molecule
- gene expression
- pet ct
- transcription factor
- metabolic syndrome
- multidrug resistant
- positron emission tomography
- high resolution
- weight loss
- genome wide
- long non coding rna
- anti inflammatory
- biofilm formation
- candida albicans
- wound healing
- genome wide identification
- glycemic control