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Ribozymes pp 45-59 | Cite as

Demonstration of a Ribozyme in Epsilon Domain of Hepatitis B Virus RNA

  • Dibyajnan Chakraborty
  • Sagarmoy GhoshEmail author
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Part of the Methods in Molecular Biology book series (MIMB, volume 2167)

Abstract

The epsilon domain of Hepatitis B virus plays a crucial role in encapsidation of viral pregenomic RNA and its partial NMR structure has been determined. However, we recently described a potassium-dependent ribonucleolytic activity associated with this region, so that a 53 nt long RNA containing the epsilon domain could release itself and cleaved other RNAs. We describe here the experimental methodologies for setting up the reactions and outline a general strategy for initial demonstration of this self-cleaving ribozyme activity.

Key words

HBV Epsilon Ribozyme Potassium 

Notes

Acknowledgments

S.G. acknowledges partial support of this work by Department of Science and Technology (Government of India) grant no. SR/FT/L-100/2004. Department of Biotechnology-BUILDER facility in University of Calcutta was used for Phosphorimager analysis. D.C. was supported by research fellowships from CSIR (9/28(664)/2006 EMR-I) and DBT (BT/01/CEIB/09/VI/10), Govt. of India.

References

  1. 1.
    Cochrane JC, Strobel SA (2008) Catalytic strategies of self-cleaving ribozymes. Acc Chem Res 41:1027–1035.  
  2. 2.
    Lilley DMJ (2011) Catalysis by the nucleolytic ribozymes. Biochem Soc Trans 39:641–646.  
  3. 3.
    Lilley DMJ (2011) Mechanisms of RNA catalysis. Philos Trans R Soc B Biol Sci 366:2910–2917.  
  4. 4.
    Fedor MJ (2009) Comparative enzymology and structural biology of RNA self-cleavage. Annu Rev Biophys 38:271–299.  
  5. 5.
    Chakraborty D, Ghosh S (2017) The epsilon motif of hepatitis B virus RNA exhibits a potassium-dependent ribonucleolytic activity. FEBS J 284:1184–1203.  
  6. 6.
    Wu HN, Lin YJ, Lin FP et al (1989) Human hepatitis delta virus RNA subfragments contain an autocleavage activity. Proc Natl Acad Sci U S A 86:1831–1835.  
  7. 7.
    Uhlenbeck OC (1987) A small catalytic oligoribonucleotide. Nature 328:596–600.  
  8. 8.
    Nesbitt S, Hegg LA, Fedor MJ (1997) An unusual pH-independent and metal-ion-independent mechanism for hairpin ribozyme catalysis. Chem Biol 4:619–630.  
  9. 9.
    Huang Z, Szostak JW (1996) A simple method for 3′-labeling of RNA. Nucleic Acids Res 24:4360–4361.  

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2021

Authors and Affiliations

  1. 1.Dr. KPC Life Sciences Pvt. Ltd.KolkataIndia
  2. 2.Department of MicrobiologyUniversity of Calcutta, University College of Science and TechnologyKolkataIndia

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