Cloning and Sequencing of Virus Inhibiting Gene Encoding an Antiviral ProteinFrom the Leaves of Pokeweed (Phytolacca americanaL.)

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Sıpahıoglu H. M., Kaya İ., Usta M., Samsun S.

Journal of Turkish Phytopathology, cilt.41, ss.19-26, 2012 (Hakemli Dergi)

Özet

Pokeweed antiviral protein (PAP) is single-chain ribosome-inactivating protein (RIP) of Phytolacca americana L. (Pokeweed) that is characterized by its ability to depurinate ribosomes. In the present study, we cloned and sequenced complete gene of pokeweed antiviral protein type 1 (PAP-I) from the summer leaves of pokeweed collected from Trabzon (Turkey) using a pair of gene specific primers based on the known N- and C-terminal nucleotide sequences of PAP gene. A product of 942 base pair was purified and inserted into pGEM-T Easy vector (Promega), downstream of the T7 promoter, and transformed into E. coli strain, JM 109. The PAP-I cistron of P. americana contained 313 amino acid residues. The DNA sequence of 942 base pairs included an open reading frame (ORF). The nucleotide sequences of PAP-I gene contained no introns and the comparison with the PAP-I sequence with the PAP isoform, showed an identity of 80-99%. Sequence analysis of PAP-I revealed that it contains a single point mutation, changing the Leucine (L) at position 273 to Phenylalanine (F) (L273F) at the putative active site.