Expression Of PKZ In Grass Carp Tissues And The Interaction Of PKZ P_Za With D(GC)₁₃ Recombinant Plasmid Visualized By AFM

The Carassius auratus PKR-like(PKZ) protein kinase,belongs to the Z-DNA binding protein family,contains an N-terminal which is composed of two Z-DNA binding domains(Za1 and Za2).To better understand the function of crucian carp PKZ,there are 3 aspects in this paper.1.The expression of PKZ gene was analysed in tissues in 28℃level by RT-PCR and it is found that there was micro-expression in tissues of grass carp at the 28℃level.When the water temperature was rapidly increased about 6℃from 28℃to 34℃to culture the grass carp for RT-PCR after 2h,it is show that the expression was increased in grass carp tissues compared with 28℃level,specially in the immune organizations of kidney and spleen.While,the same results we found in the tissues of grass carp which was induced by Poly I:C after 2d by RT-PCR.2.The CaPKZ C teminal domain whose sequence length is about 1kb had been cloned.And the C teminal fusion peptide was expressed in BL21PlysS prokaryotic expression system and then purified by His affinity chromatography. Gel mobility shift assay revealed that the fusion protein is about 53KD.The expression and purification of PKZ C teminal is for the function study of PKZ in future.3.Three kinds of recombinant Za peptides(P_Za1,P_Za2,P_Za1Za2) were expressed by a prokaryotic expression system and then purified by affinity chromatography. Interaction of the three kinds of Za proteins of CaPKR-like(PKZ) with pMD18-T and d(GC)₁₃ recombinant plasmid were visualized by atomic force microscopy. The results indicated that none of P_Za1,P_Za2 and P_Za1Za2 could bind to pMD18-T. But P_Za1Za2,rather than P_Za1 and P_Za2,could bind to d(GC)₁₃ recombinant plasmid.