Production of native protein by using Synechocystis sp. PCC6803 DnaB mini-intein in Escherichia coli.
To directly express native recombinant proteins in Escherichia coli, a new expression vector pSB was constructed using Ssp DnaB mini-intein. Using the vector, native proteins could be produced with the help of C-terminal self-cleavage of the intein. In this study, we cloned hIFNalpha-4 gene into pSB and used E. coli strain Origami B (DE3) as the host. Expression experiments were carried out both in Shake flasks and a 5 L bioreactor. The results indicated hIFNalpha-4 could be expressed in the form of soluble protein with correct folding in E. coli. The maximal hIFNalpha-4 content was 21.7% of total protein, and the antiviral activity of the protein was 1.2x10(8 )IU mg(-1). Overall, good effects were achieved with this system. This intein-mediated protein expression system opens up a useful method for production of native recombinant protein in E. coli.
Yan
Shou-Sheng
SS
School of Pharmaceutical Hunan College of TCM, Changsha, Hunan, China. yanshousheng@163.com
Yan
Juan
J
Shi
Gang
G
Xu
Qi
Q
Chen
Shou-Chun
SC
Tian
Yu-Wang
YW
D000998:Antiviral Agents / Q000138:chemical synthesis; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D006801:Humans; D047668:Inteins / Q000235:genetics*; D007370:Interferon Type I / Q000235:genetics / Q000494:pharmacology; D016898:Interferon-alpha; D011994:Recombinant Proteins / Q000235:genetics*; D046939:Synechocystis / Q000737:chemistry*; D014721:Vesicular stomatitis Indiana virus / Q000187:drug effects
D000998:Antiviral Agents; D007370:Interferon Type I; D016898:Interferon-alpha; D011994:Recombinant Proteins; C068567:interferon alpha 4A