Cloning and Sequencing of Virus Inhibiting Gene Encoding an Antiviral Protein From the Leaves of Pokeweed (Phytolacca Americana L.)

Authors : Hikmet Murat SİPAHİOĞLU, İlhan KAYA, Mustafa USTA, Sibel SAMSUN KIZILDAĞ

Pages : 19-26

View : 17 | Download : 13

Publication Date : 2014-05-28

Article Type : Research Paper

Abstract :800x600 Normal 0 21 false false false TR X-NONE X-NONE MicrosoftInternetExplorer4 st1\:*{behavior:urlinsert ignore into journalissuearticles values(#ieooui); } /* Style Definitions */ table.MsoNormalTable {mso-style-name:`Normal Tablo`; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:``; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:`Times New Roman`,`serif`;} Pokeweed antiviral protein insert ignore into journalissuearticles values(PAP); is single-chain ribosome-inactivating protein insert ignore into journalissuearticles values(RIP); of Phytolacca americana L. insert ignore into journalissuearticles values(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 insert ignore into journalissuearticles values(PAP-I); from the summer leaves of pokeweed collected from Trabzon insert ignore into journalissuearticles values(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 insert ignore into journalissuearticles values(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 insert ignore into journalissuearticles values(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 insert ignore into journalissuearticles values(L); at position 273 to Phenylalanine insert ignore into journalissuearticles values(F); insert ignore into journalissuearticles values(L273F); at the putative active site.
Keywords : antiviral proteins, characterization, cloning, PAP I gene, phytolacca americana