Gene Cloning and Bioinformatics Analysis of Nitrite Reductase from Bacillus cereus LJ01

To study the enzymatic properties of nitrite reductase(NiR)in Bacillus cereus LJ01 and obtain highly expressed genetic engineering bacteria,the NiR sequence of B. cereus LJ01 was analyzed by bioinformatics. Then the NiR gene was cloned into expression vectors pET-28a(+)and pET-32a(+),and the genetic engineering bacteria pET-28a(+)-nir-BL21 and pET-32a(+)-nir-BL21 were constructed. Afterwards,the effects of different induction temperatures on the expression of recombinant NiR was studied. The results showed that the theoretical molecular weight of the protein encoded by NiR was about 60 kDa,the theoretical pI was 5.47,the secondary structure was mainly alpha-helix and irregular curl,and it was a hydrophilic protein without transmembrane structure. With the increasing of induction temperature,the expression of recombinant NiR decreased gradually,and the highest expression of recombinant NiR was found at the induction temperature of 16℃. At the same induction temperature,the expression of NiR in pET-28a(+)-nir-BL21 was significantly higher than that in pET-32a(+)-nir-BL21.Therefore,pET-28a(+)-nir-BL21 was selected as the genetic engineering bacteria. This study cloned NiR from obtained B. cereus LJ01 and obtained pET-28a(+)-nir-BL21 as engineering bacteria,which laid a foundation for the study of the physical and chemical properties of the recombinant NiR and the heterologous expression of NiR in foodborne Bacillus spp.