Construction of a New Genetic Engineering Bacterium for Preparation of Superoxide Dismutase with High Productivity

Abstract:

A new Mn-SOD gene encoding 202 amino acids was cloned from genomic DNA of Bacillus subtilis ATCC 9372 for construction of a genetic engineering bacterium to produce SOD. Its phylogenetic relationships with other Bacillus spp. revealed that this predicted protein is most closely related to B. atrophaeus NRS-213 (AY197616) and B. subtilis 168. This gene was inserted into expression plasmid pET28a and first successfully expressed in E. coli BL21. The SOD was expressed accounted for approximately 45.6% of total bacterial protein. The activity of the SOD was 2553.211 U/mg, the enzyme showed maximum activity at about pH 8.0 and relatively stable from pH 6.0 to 11.0. This SOD had a good thermal stability with >75% retaining of the relative enzymatic activity after incubation at 50 °C for 90 min. This study demonstrated that a new genetic engineering bacterium to produce SOD with high productivity has been successfully constructed.

• Periodical: Advanced Materials Research (Volumes 314-316)
• Pages: 1973-1976
• DOI: https://doi.org/10.4028/www.scientific.net/AMR.314-316.1973
• Online since: August 2011
• Authors: Yi Zhao, Yang Li, Ren Qiang Li
• Keywords: B. Subtilis ATCC 9372, Clone, Expression, Genetic Engineering Bacterium, Mn-Sod

References

[1] P. Liu, H.E. Ewis, Y.J. Huang, C.D. Lu, P.C. Tai and I.T. Weber: Acta Crystallogr Sect F Struct Biol Cryst Commun Vol. 63(2007), p.1003

DOI: 10.1107/s1744309107054127

[2] R. Pipe, C. Porte and D. Livingstone: Fish Shellfish Immunol Vol. 3(1993), p.221

[3] I. Fridovich: Annu Rev Biochem Vol. 64(1995), p.97

[4] G.A. Gómez-Anduro, C.V. Barillas-Mury, A.B. Peregrino-Uriarte, L. Gupta, T. Gollas-Galván, J. Hernández-López and G. Yepiz-Plascencia: Dev Comp Immunol Vol. 30(2006), p.893

[5] I. Fridovich: Adv Enzymol Relat Areas Mol Biol Vol. 41(1986), p.35

[6] H.D. Youn, H. Youn, J.W. Lee, Y.I. Yim, J.K. Lee, Y.C. Hah and S.O. Kang: Arch Biochem Biophys Vol. 334(1996), p.341

[7] F.J. Kim, H.P. Kim, Y.C. Hah and J.H. Roe: Eur J Biochem Vol. 241(1996),p.178

[8] M. Osawa, F. Yamakura, M. Mihara, Y. Okubo, K. Yamada and B.Y. Hiraoka: Biochim Biophys Acta Vol. 1804(2010), p.1775

[9] T. Inaoka, Y. Mastumura and T. Tsuchido: J Bacteriol Vol. 181(1999), P. (1939)

[10] T. Inaoka, Y. Matsumura and T. Tsuchido: J Bacteriol Vol. 180(1998) , P. 3697

[11] Y.Z. He, K.Q. Fan, C.J. Jia, Z.J. Wang, W.B. Pan, L. Huang, K.Q. Yang and Z.Y. Dong: Appl Microbiol Biotechnol Vol. 75(2007), p.367

[12] T. Hunter, J.V. Bannister and G.J. Hunter: Eur J Biochem Vol. 269 (2002), p.5137

[13] S. Steffen, K. Silke and Gunnar W: Plant Cell Physiol Vol. 35(1994), p.859

[14] T. Nakanishi, H. Inoue and M. Kitamura: J Biochem Vol. 133(2003) , p.387