Skip to main content
SpringerLink
Log in

Purification and characterization of a hyperthermostable Mn-superoxide dismutase from Thermus thermophilus HB27

  • Original Paper
  • Published:
Extremophiles Aims and scope Submit manuscript

Abstract

Thermostable Mn-dependent catalases are promising enzymes in biotechnological applications. In the present study, a Mn-containing superoxide dismutase of the hyperthermophilic Thermus thermophilus HB27 had been purified and characterized by a two-stage ultrafiltration process after being expressed in E. coli. The enzyme was highly stable at 90°C and retained 57% activity after heat treatment at 100°C for 1 h. The native form of the enzyme was determined as a homotetramer by analytical size exclusion chromatography and sodium dodecyl sulfate–polyacrylamide gel electrophoresis. The final purified enzyme had an isoelectric point of 6.2 and a high α-helical content of 70%, consistent with the theoretical values. This showed that the purified SOD folded with a reasonable secondary structure.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

Abbreviations

Mn-SOD:

Manganese superoxide dismutase

CD:

Circular dichroism

IEF:

Isoelectric focusing

SDS–PAGE:

Sodium dodecyl sulfate–polyacrylamide gel electrophoresis

IgG:

Immunoglobulin G

BSA:

Bovine serum albumin

PES:

Polyethersulfone

MWCO:

Molecular weight cut-off

RC:

Regenerated cellulose

PCR:

Polymerase chain reaction

pI :

Isoelectric point

E. coli :

Escherichia coli

References

  • Chang QL, Chen JY, Zhang XF, Zhao NM (1997) Effect of the cosolvent type on the extraction of α-amylase with reversed micelles: circular dichroism study. Enzyme Microb Technol 20:87–92

    Article  CAS  Google Scholar 

  • Gu HZ, Liu LQ, Zhao QS (2006) A comparison between two methods for determination of SOD activity. Chin Pharm Biotechnol 13:377–379

    CAS  Google Scholar 

  • Henne A, Brüggemann H, Raasch C, Wiezer A, Hartsch T, Liesegang H, Johann A, Lienard T, Gohl O, Martinez-Arias R, Jacobi C, Starkuviene V, Schlenczeck S, Dencker S, Huber R, Klenk HP, Kramer W, Merkl R, Gottschalk G, Fritz HJ (2004) The genome sequence of the extreme thermophile Thermus thermophilus. Nat Biotechnol 22:547–553

    Article  PubMed  CAS  Google Scholar 

  • Holdom MD, Hay RJ, Hamilton AJ (1995) Purification, N-terminal amino acid sequence and partial characterization of a Cu, Zn superoxide dismutase from the pathogenic fungus Aspergillus fumigatus. Free Radical Res 22:519–531

    Article  CAS  Google Scholar 

  • Holdom MD, Hay RJ, Hamilton AJ (1996) The Cu, Zn superoxide dismutases of Aspergillus flavus, Aspergillus niger, Aspergillus nidulans, and Aspergillus terreus: purification and biochemical comparison with the Aspergillus fumigatus Cu, Zn superoxide dismutase. Infect Immun 64:3326–3332

    PubMed  CAS  Google Scholar 

  • Koyama Y, Hoshino T, Tomizuka N, Furukawa K (1986) Genetic transformation of the extreme thermophile Thermus thermophilus and of other Thermus spp. J Bacteriol 166:338–340

    PubMed  CAS  Google Scholar 

  • Kumar S, Sahoo R, Ahuja PS (2006) Isozyme of autoclavable superoxide dismutase (SOD), a process for the identification and extraction of the SOD and use of the said SOD in cosmetic, food, and pharmaceutical compositions. US Patent 7037697 B2

  • Li DC, Guo J, Li YL, Lu J (2005) A thermostable manganese-containing superoxide dismutase from the thermophilic Thermomyces lanuginosus. Extremophiles 9:1–6

    Article  PubMed  Google Scholar 

  • Lim JH, Yu YG, Han YS, Cho S, Ahn BY, Kim SH, Cho Y (1997) The crystal structure of an Fe-superoxide dismutase from the hyperthermophile Aquifex pyrophilus at 1.9 A resolution: structural basis for thermostability. J Mol Biol 270:259–274

    Article  PubMed  CAS  Google Scholar 

  • Liu JG, Lu JR, Zhao XB, Lu JR, Cui ZF (2007) Separation of glucose oxidase and catalase using ultrafiltration with 300-kDa polyethersulfone membranes. J Membr Sci 299:222–228

    Article  CAS  Google Scholar 

  • Liu JG, Yang J, Xu H, Lu JR, Cui ZF (2010) A new membrane based process to isolate immunoglobulin from chicken egg yolk. Food Chem 122:747–752

    Article  CAS  Google Scholar 

  • Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275

    PubMed  CAS  Google Scholar 

  • McCord JM, Fridovich I (1969) Superoxide dismutase: an enzymic function for erythrocuprein (hemocuprein). J Biol Chem 244:6049–6055

    PubMed  CAS  Google Scholar 

  • Michalski WP (1996) Chromatographic and electrophoretic methods for analysis of superoxide dismutases. J Chromatogr B 684:59–75

    Article  CAS  Google Scholar 

  • Neuilly-sur-Seine GK, Atony BJ, Deuil-la-Barre GL (1978) Protecting skin and hair with cosmetic compositions containing superoxide dismutase. US Patent 4129644

  • Sharma NC, Sahi SV, Jain JC (2005) Sesbania drummondii cell cultures: ICP-MS determination of the accumulation of Pb and Cu. Microchem J 81:163–169

    Article  CAS  Google Scholar 

  • Whittaker MM, Whittaker JW (2000) Recombinant superoxide dismutase from a hyperthermophilic archaeon, Pyrobaculum aerophilum. J Bio Inorg Chem 5:402–408

    CAS  Google Scholar 

  • Xu SH, Hu H, Li YP (2001) Improvement of superoxide dismutase activity test of pyrogallol autooxidation method. Chin Chem Bioeng 8:516–519

    Google Scholar 

  • Yamano S, Maruyama T (1999) An azide-insensitive superoxide dismutase from a hyperthermophilic archaeon, Sulfolobus solfataricus. J Biochem 125:186–193

    PubMed  CAS  Google Scholar 

  • Yamano S, Sako Y, Nomura N, Maruyama T (1999) A cambialistic SOD in a strictly aerobic hyperthermophilic archaeon, Aeropyrum pernix. J Biochem 126:218–225

    PubMed  CAS  Google Scholar 

  • Yang JT, Wu CC, Martinez HM (1986) Poly (L-lysine) at different pH and temperature. Methods Enzymol 130:208–271

    Article  PubMed  CAS  Google Scholar 

  • Zhang GH, Ge HB, Li QY, Zhang XY (2004) Role of SOD in protection strawberry leaves from photo-inhibition damage. Chin J Fruit Sci 21:328–330

    Google Scholar 

Download references

Acknowledgments

The authors are grateful for the financial support of the Doctoral Foundation of Shandong Province (No. 2008BS02018).

Author information

Authors and Affiliations

  1. Center for Bioengineering and Biotechnology, China University of Petroleum (East China), Qingdao, 266555, People’s Republic of China

    Jianguo Liu, Mengmeng Yin & Hu Zhu

  2. School of Physics, Manchester University, Manchester, M60 1QD, UK

    Jianren Lu

  3. Department of Engineering Science, University of Oxford, Oxford, OX1 3PJ, UK

    Zhanfeng Cui

Authors
  1. Jianguo Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mengmeng Yin
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Hu Zhu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jianren Lu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Zhanfeng Cui
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Hu Zhu.

Additional information

Communicated by A. Driessen.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Liu, J., Yin, M., Zhu, H. et al. Purification and characterization of a hyperthermostable Mn-superoxide dismutase from Thermus thermophilus HB27. Extremophiles 15, 221–226 (2011). https://doi.org/10.1007/s00792-010-0350-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00792-010-0350-3

Keywords

Search

Navigation