Snake venoms represent a danger to human health, but also a gold mine of bioactive proteins that can be harnessed for drug discovery purposes. The evolution of snakes and their venom has been studied for decades, particularly via traditional morphological and basic genetic methods alongside venom proteomics. However, while the field of genomics has matured rapidly over the past 2 decades, owing to the development of next-generation sequencing technologies, snake genomics remains in its infancy. Here, we provide an overview of the state of the art in snake genomics and discuss its potential implications for studying venom evolution and toxinology. On the basis of current knowledge, gene duplication and positive selection are key mechanisms in the neofunctionalization of snake venom proteins. This makes snake venoms important evolutionary drivers that explain the remarkable venom diversification and adaptive variation observed in these reptiles. Gene duplication and neofunctionalization have also generated a large number of repeat sequences in snake genomes that pose a significant challenge to DNA sequencing, resulting in the need for substantial computational resources and longer sequencing read length for high-quality genome assembly. Fortunately, owing to constantly improving sequencing technologies and computational tools, we are now able to explore the molecular mechanisms of snake venom evolution in unprecedented detail. Such novel insights have the potential to affect the design and development of antivenoms and possibly other drugs, as well as provide new fundamental knowledge on snake biology and evolution.

Disintegrin-like/cysteine-rich (DC) proteins have long been regarded just as products of proteolysis of P-III snake venom metalloproteinases (SVMPs). However, here we demonstrate that a DC protein from the venom of Vipera ammodytes (Vaa; nose-horned viper), VaaMPIII-3, is encoded per se by a P-III SVMP-like gene that has a deletion in the region of the catalytic metalloproteinase domain and in part of the non-catalytic disintegrin-like domain. In this way, we justify the proposal of the introduction of a new subclass P-IIIe of SVMP-derived DC proteins. We purified VaaMPIII-3 from the venom of Vaa in a series of chromatographic steps. A covalent chromatography step based on thiol-disulphide exchange revealed that VaaMPIII-3 contains an unpaired Cys residue. This was demonstrated to be Cys6 in about 90% and Cys19 in about 10% of the VaaMPIII-3 molecules. We further constructed a three-dimensional homology model of VaaMPIII-3. From this model, it is evident that both Cys6 and Cys19 can pair with Cys26, which suggests that the intramolecular thiol-disulphide exchange has a regulatory function. VaaMPIII-3 is an acidic 21-kDa monomeric glycoprotein that exists in at least six N-glycoforms, with isoelectric points ranging from pH 4.5 to 5.1. Consistent with the presence of an integrin-binding motif in its sequence, SECD, VaaMPIII-3 inhibited collagen-induced platelet aggregation. It also inhibited ADP- and arachidonic-acid-induced platelet aggregation, but not ristocetin-induced platelet agglutination and the blood coagulation cascade.

We present a chromosome-level genome assembly of the western nose-horned viper (Vipera ammodytes ammodytes), the most medically important viper in Europe. Using PacBio Sequel and Illumina HiSeq X Ten sequencing, we generated ∼270 Gb of data, achieving ∼131× coverage of the genome. The final assembly spans 1.55 Gb with a contig N50 of 45.9 Mb and a scaffold N50 of 210 Mb, anchored into 18 pseudo-chromosomes. Completeness was supported by recovery of 97.1% of Vertebrata BUSCOs. A total of 20,775 protein-coding genes were predicted, of which 96.6% were functionally annotated. Repetitive sequences accounted for 53.75% of the genome, dominated by LINEs (41.87%) and LTRs (14.35%). We identified 112 venom-related genes across 15 families, with expansions in SVMPs, Snaclecs, sPLA₂s, SPIs, and SVSPs, together comprising 62.5% of the venom repertoire. Chemosensory genes were also expanded, including 448 olfactory receptors, 72 taste receptors, and 29 vomeronasal receptors. This assembly represents the most complete genome resource for a true viper to date and provides a key resource for investigating venom evolution, chemosensory adaptation, and comparative snake genomics.