Cloning and Prokaryotic Expression and Structure Prediction of Prx II the Differentially Expressed Gene under AFB1 Stress
The gene, prx II, in mice liver was found to be up-regulated under the stress of AFB1 in our previous study. In this study, the total RNA from the mice liver was extracted, and followed by the synthesis of cDNA with the RNA as template by the method of reverse transcription. Then, prx II gene fragment was amplified by PCR with the cDNA as template. After double-digestion and ligation reaction, the prokaryotic expression vector pET28a (+)-prx II was successfully constructed and was transformed into E. coli BL21 (DE3). The protein- Prx II was successfully expressed when induced by IPTG, and finally, Prx II was purified by Ni2+-NTA affinity chromatography. The molecule structure of Prx II, including its signal peptide, hydrophobicity, and its secondary and tertiary structure, was predicted by bioinformatics analysis. The results showed that no signal peptide was predicted in the molecule of Prx II; Five hydrohpobic domains were predicted in the protein molecule, and the average predictive value for its hydrophobicity was -0.151; There were 35% α-helix (69 residues) and 21% β-pleated sheet (42 residues) in the molecule; The tertiary structure of the protein was constituted by seven α-helices and seven β-pleated sheets.
Zhong Cao, Yinghe He, Lixian Sun and Xueqiang Cao
Z. H. Zhuang et al., "Cloning and Prokaryotic Expression and Structure Prediction of Prx II the Differentially Expressed Gene under AFB1 Stress", Advanced Materials Research, Vols. 236-238, pp. 1078-1082, 2011