Premium
ATP and the cap analog, m7GTP, bind competitively to the active site of Pokeweed Antiviral Protein (PAP)
Author(s) -
Friedland Diana Elaine,
Goss Dixie J.,
Baldwin Amy E.
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a650-b
Subject(s) - binding site , biochemistry , nucleotide , ribosome inactivating protein , a site , ribosome , biology , ribosomal protein , ricin , ribosomal binding site , chemistry , rna , toxin , gene
Ribosome‐inactivating proteins (RIPs) are naturally occurring cytotoxic agents and are found in numerous plant, fungi, and bacterial species. They inhibit protein synthesis by depurinating the conserved sarcin/ricin loop (SRL) of the large ribosomal RNA [1]. PAP has been shown to bind to the cap analogs and depurinate sites downstream of the cap. The mechanism of selection of mRNA remains unknown. Modeling studies have suggested that the cap binding interaction and depurination activity occur at the same site on PAP. However, there is no binding pocket homology between PAP's active site and the binding pocket of other cap binding proteins. Here we report quantitative results of steady‐state fluorescence studies of the effects of nucleotides on PAP binding to mRNA cap. ATP and other nucleotides show competitive binding with m7GTP for PAP. These data strongly suggest PAP's cap binding site and enzymatic site are the same. All nucleotides bind PAP in the absence of cap (1.5 – 2.0 times weaker than cap). PAP binds cap with about three‐fold lower affinity that wheat germ cap binding protein, eIFiso4E. However, the concentration of PAP is probably much higher than eIFiso4E allowing for competition with cellular cap binding proteins. The effects of other initiation factors on PAP cap binding will be reported. This work was supported in part by a Research Opportunity Award (NSF MCB 0413982 and a CUNY Collaborative Incentive Research Grant 80209‐07‐12).