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3 Biotech

, 7:201 | Cite as

Recombinant production of Aspergillus Flavus uricase and investigation of its thermal stability in the presence of raffinose and lactose

  • Mehdi Imani
  • Serveh Shahmohamadnejad
Original Article

Abstract

Aspergillus flavus uricase (Rasburicase) with a molecular mass of 135 kDa is currently used for the treatment of gout and hyperuricemia occurring in tumor lysis syndrome. To characterize the effects of raffinose and lactose osmolytes on the uricase structure, its coding sequence was cloned, expressed in E. coli BL21, and purified by Ni–NTA agarose affinity chromatography. Thermal inactivation studies at 40 °C showed that nearly 15% of UOX activity was preserved, while the presence of raffinose and lactose reduced its activity to 35 and 45% of its original activity, respectively. Investigation of UOX thermal stability at 40 °C in the course of time showed that the enzyme relatively lost almost 60% of its original activity after 40 min, whereas more than 50% of UOX activity is preserved in the presence of lactose. Estimation of thermal inactivation rate constant, k in, showed that the UOX k in and UOX k in in the presence of raffinose was unchanged (0.018 min−1), whereas for the presence of lactose, it was 0.015 min−1. Half-life and T m analysis showed that UOX half-life is almost 38 min and addition of raffinose did not change the half-life, whereas the presence of lactose had remarkable impact on UOX half-life (46 min). The presence of raffinose increased UOX T m to a lesser extent, whereas lactose notably enhanced the T m from 27 to 37 °C. Overall, our findings show that lactose has protective effects on UOX stability, while for raffinose, it is relatively compromised.

Keywords

Uricase Osmolyte Thermal stability Recombinant Aspergillus flavus 

Notes

Acknowledgements

The authors would like to thank the research council of Urmia University for financial support. Technical support from Mr. Ashkan Basirnia and Mrs. Razieh Pak-Tarmani is highly appreciated, as well.

Compliance with ethical standards

Conflict of interest

The authors report no conflicts of interest.

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Copyright information

© Springer-Verlag GmbH Germany 2017

Authors and Affiliations

  1. 1.Department of Basic Sciences, Faculty of Veterinary MedicineUrmia UniversityUrmiaIran
  2. 2.Department of Cellular and Molecular Biotechnology, Institute of BiotechnologyUrmia UniversityUrmiaIran

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