Purification, characterization and cloning of antiviral/ribosome inactivating protein from Amaranthus tricolor leaves |
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Authors: | Roy Sribash Sadhana P Begum Mehbuba Kumar Sushil Lodha M L Kapoor H C |
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Affiliation: | Division of Biochemistry, Indian Agricultural Research Institute, New Delhi 110012, India. |
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Abstract: | An antiviral protein (AVP), imparting high level of resistance against sunnhemp rosette virus (SRV) was purified from the dried leaves of Amaranthus tricolor. The purified protein (AAP-27) exhibited approximately 98% inhibition of local lesion formation at a concentration range of approximately 30 microg ml(-1). The protein was found to be highly basic glycoprotein monomer (pI approximately 9.8) of Mr 27 kDa, with neutral sugar content of 4%. The purified protein exhibited N-glycosidase and RNase activities. We have also isolated full-length cDNA clone, encoding this protein designated as A. tricolor antiviral protein-1 (AAP-1). Two primers, one designed on the basis of N-terminal sequence of the purified protein and the other from the conserved active peptides of other AVPs/RIPs were used for PCR amplification of double stranded cDNA, isolated from the leaves of A. tricolor. The amplified fragment was used as a probe for library screening. The isolated full-length cDNA consisted of 1058 nucleotides with an open reading frame encoding a polypeptide of 297 amino acids. The deduced amino acid sequence of AAP-1 has a putative active domain conserved in other AVPs/RIPs and shows varying homology to the RIPs from other plant species. |
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Keywords: | Amaranthus tricolor Antiviral protein Protein purification Basic glycoprotein N-glycosidase activity RNase activity cDNA |
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