Efficient fusion expression of G13 domain derived from granulysin in Escherichia coli.
- Author:
Xiaoqiang LIU
1
;
Xiangdong ZHA
;
Yazhong XIAO
;
Jinhuan YANG
;
Nengshu LI
Author Information
1. Key Laboratory of Ecological Engineering and Biotechnology of Anhui Province, School of Life Science, Anhui University, Hefei 230039, China.
- Publication Type:Journal Article
- MeSH:
Anti-Infective Agents;
metabolism;
Antigens, Differentiation, T-Lymphocyte;
genetics;
Cyanogen Bromide;
pharmacology;
Escherichia coli;
genetics;
metabolism;
GTP-Binding Protein alpha Subunits, G12-G13;
biosynthesis;
genetics;
Inclusion Bodies;
metabolism;
Protein Structure, Tertiary;
genetics;
Recombinant Fusion Proteins;
biosynthesis;
genetics;
Thioredoxins;
genetics;
Transfection
- From:
Chinese Journal of Biotechnology
2009;25(2):235-241
- CountryChina
- Language:Chinese
-
Abstract:
The G13 domain derived from granulysin shows high antimicrobial activities against Gram-positive and Gram-negative bacteria but does not lyse Jurkat cells or liposomes. To explore a new approach for high expression of the G13 domain, we fused the sequence encoding G13 to thioredoxin (Trx) gene to construct the recombinant expression vector (pThioHisA-G13). A cyanogen bromide (CNBr) cleavage site was introduced between the Trx and G13 to facilitate final release of the recombinant G13. The recombinant expression vector, pThioHisA-G13, was transformed into E. coli BL21 (DE3). Upon induction by IPTG Trx-G13 fusion protein was expressed and took the form of inclusion bodies counting 58% (W/W) of total cellular proteins. The inclusion body was solved by urea (8 mol/L) and then cleaved by CNBr. We purified the recombinant peptide G13 by one-step cation exchange chromatography. Results of agarose diffuse assay analysis indicated that the recombinant G13 exhibited antibacterial activity. The procedure described in this study will provide a reliable and simple method for highly efficient production of some cationic antimicrobial peptides.
