Molecular cloning and functional identification of a ribosome inactivating/antiviral protein from leaves of post-flowering stage of Celosia cristata and its expression in E. coli

Mehbuba Begam, Sushil Kumar, Sribash Roy, James J. Campanella, H. C. Kapoor

Research output: Contribution to journalArticle

15 Scopus citations

Abstract

A full-length cDNA clone, encoding a ribosome inactivating/antiviral protein (RIP/AVP) was isolated from the cDNA library of post-flowering stage of Celosia cristata leaves. The full-length cDNA consisted of 1015 nucleotides, with an open reading frame encoding 283 amino acids. The deduced amino acid sequence had a putative active site domain conserved in other ribosome inactivating/antiviral proteins (RIPs/AVPs). The coding region of the cDNA was amplified by polymerase chain reaction (PCR), cloned and expressed in Escherichia coli as recombinant protein of 72 kDa. The expressed fusion product was confirmed by Western analysis and purification by affinity chromatography. Both the recombinant protein (reCCP-27) and purified expressed protein (eCCP-27) inhibited translation in rabbit reticulocytes showing IC50 values at 95 ng and 45 ng, respectively. The native purified nCCP-27 has IC50 at 25 ng. The purified product also showed N-glycosidase activity towards tobacco ribosomes and antiviral activity towards tobacco mosaic virus (TMV) and sunnhemp rosette virus (SRV).

Original languageEnglish
Pages (from-to)2441-2449
Number of pages9
JournalPhytochemistry
Volume67
Issue number22
DOIs
StatePublished - 1 Nov 2006

Keywords

  • Antiviral
  • Celosia cristata
  • Expression
  • Ribosome inactivating
  • cDNA

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