Abstract: In order to study the role and function of the nuclease AtCaN2 gene in plant growth and development,the AtCaN2 gene was cloned from Arabidopsis thaliana. Bioinformatics analysis,subcellular localization analysis,protein induction,expression,purification and functional were carried out to study this gene. The results showed that the isoelectric point of AtCaN2 protein was 8.45,the chemical formula was C₁₁₁₆H₁₇₈₈N₃₁₀O₃₃₅S₃,which contained 3 552 atoms and it was a hydrophilic protein; AtCaN2 had high homology with Camelina sativa,the homology was 87.89%,and it had low homology with Setaria italic, Sorghum bicolor and Zea mays,the homology was 56.42%,57.35% and 56.47% respectively,among the amino acid sequences,there was a highly conserved D-X-D sequence; evolutionary tree analysis showed that AtCaN2 was closely related to C.sativa,but far from S.italic and Z.mays,whis were consistent with the results of amino acid sequence alignment; subcellular localization results showed that it was located in cytoplasm,which was consistent with the predicted results; the His-AtCaN2 fusion protein was induced and purified to be about 35 ku in size; Western Blot analysis showed that the fusion protein was correctly translated and expressed without structural breakage or degradation; the fusion protein of His-AtCaN2(M)was expressed in small amount and large amount of induction,and the single band of His-AtCaN2(M)was obtained after purification; enzyme activity analysis showed that His-AtCaN2 fusion protein had the ability to degrade the nucleic acid substrate,showing high activity,while the His-AtCaN2(M)point mutation fusion protein had no ability to degrade the nucleic acid substrate,and the substrate was basically not degraded.The results of this study provide a basis for further study of the related mechanism of AtCaN2 in plants.

Key words: Arabidopsis thaliana, Nuclease, AtCaN2, Subcellular localization, Prokaryotic expression, Enzyme activity

CLC Number: 

• Q78

• S55