Purification, characterization and cloning of antiviral/ribosome inactivating protein from Amaranthus tricolor leaves.
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.
Roy
Sribash
S
Division of Biochemistry, Indian Agricultural Research Institute, New Delhi 110012, India.
Sadhana
P
P
Begum
Mehbuba
M
Kumar
Sushil
S
Lodha
M L
ML
Kapoor
H C
HC
D027721:Amaranthus / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000998:Antiviral Agents / Q000737:chemistry / Q000302:isolation & purification; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D003609:Dactinomycin / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006026:Glycoside Hydrolases / Q000378:metabolism; D007525:Isoelectric Focusing; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018515:Plant Leaves / Q000235:genetics* / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction; D012260:Ribonucleases / Q000378:metabolism; D017422:Sequence Analysis, DNA; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid
D000998:Antiviral Agents; D018076:DNA, Complementary; D010940:Plant Proteins; D003609:Dactinomycin; D012260:Ribonucleases; D006026:Glycoside Hydrolases