High-throughput proteomics and in vitro functional characterization of the 26 medically most important elapids and vipers from sub-Saharan Africa.

Venomous snakes are important parts of the ecosystem, and their behavior and evolution have been shaped by their surrounding environments over the eons. This is reflected in their venoms, which are typically highly adapted for their biological niche, including their diet and defense mechanisms for deterring predators. Sub-Saharan Africa is rich in venomous snake species, of which many are dangerous to humans due to the high toxicity of their venoms and their ability to effectively deliver large amounts of venom into their victims via their bite. In this study, the venoms of 26 of sub-Saharan Africa's medically most relevant elapid and viper species were subjected to parallelized toxicovenomics analysis. The analysis included venom proteomics and in vitro functional characterization of whole venom toxicities, enabling a robust comparison of venom profiles between species. The data presented here corroborate previous studies and provide biochemical details for the clinical manifestations observed in envenomings by the 26 snake species. Moreover, two new venom proteomes (Naja anchietae and Echis leucogaster) are presented here for the first time. Combined, the presented data can help shine light on snake venom evolutionary trends and possibly be used to further improve or develop novel antivenoms.

Epitope mapping and characterization of 4-hydroxy-2-nonenal modified-human serum albumin using two different polyclonal antibodies.

Lipids are susceptible to damage by reactive oxygen species, and from lipid oxidation reactions many short chain lipid peroxidation products can be formed. 4-Hydroxy-2-nonenal (HNE) is one of the most abundant and cytotoxic lipid oxidation products and is known to form covalent adducts with nucleophilic amino acids of proteins. HNE-modified proteins have value as biomarkers and can be detected by antibody-based techniques, but most commercially available antibodies were raised against HNE-keyhole limpet hemocyanin. We used HNE-treated human serum albumin (HSA) to raise sheep antiserum and report for the first time the use of covalently modified peptide arrays to assess epitope binding of antibodies (Abs). Peptide arrays covering the sequence of HSA and treated post peptide synthesis with HNE were used to compare the different binding patterns of a commercial polyclonal antibody (pAb) raised against HNE-treated KLH and an in-house anti-HNE enriched pAb. The results were correlated with analysis of HNE-modified HSA by high-resolution tandem mass spectrometry. Both anti-HNE pAbs were found to bind strongly to eight common peptides on the HNE-treated HSA membranes, suggesting that HNE adducts per se induced an immune response in both cases even though different immunogens were used. Both antibodies bound with the highest affinity to the peptide 365DPHECYAKVFDEFKPLV381, which contains K378 and was also shown to be modified by the mass spectrometry analysis. Overall, the commercial anti-HNE pAb showed better specificity, recognizing nine out of the eleven adducts found by MS/MS, while the in-house enriched pAb only recognizes six. Nevertheless, the in-house pAb recognized specific peptides that were not recognized by the commercial pAb, which suggests the presence of clones uniquely specific to HNE adducts on HSA.