Exploring the venom proteome of the African puff adder, Bitis arietans, using a combinatorial peptide ligand library approach at different pHs

We report the 2DE-based proteomic characterization of the venom of the medically important African puff adder, Bitis arietans, after prefractionation by incubation with a solid-phase combinatorial hexapeptide ligand library (CPLL) at three different pH values. This approach yielded partially overlapping yet clearly distinct sets of proteins. The B. arietans venom proteome, merged from the four sets of proteins comprises at least 43 distinct proteins from 9 toxin families. In line with a previous reverse-phase HPLC-based venomic characterization on the same species, SVMPs, serine proteinases, C-type lectin-like proteins, and to a minor extent PLA2, disintegrin bitistatin, and cystatin, comprise the major toxins in the venom of B. arietans. However, the 2D-CPLL approach employed here identified both a significantly higher (about double) number of proteins than a previous venomic approach, and many very minor components barely, or not at all, detectable in the 2DE separation of whole venom. 30 proteins from the CPLL-merged venom proteome matched some of the 63 toxin clusters generated by sequencing one thousand randomly selected venom gland cDNA library clones of the same species. The low (47%) concordance between transcriptome and proteome may be interpreted in terms of intraspecific venom variation. Comparison of the reverse-phase HPLC separations of the venom proteins of B. arietans from Ghana and Nigeria supports this view.

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