Unlocking the toxic arsenal of bioactive proteins from snake venoms for biotherapeutic applications

Weiqiao Rao*

*Corresponding author for this work

Research output: Book/ReportPh.D. thesis

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Abstract

The threat of snakebites has been declared a global health emergency. According to WHO estimates, around 5.4 million people are bitten by snakes every year, resulting in up to 138,000 deaths and 400,000 people with permanent disabilities. Venomous snakes use venom as a weapon to capture their prey and for defensive purposes. Antivenom is the definitive treatment for venomous snakebites, and the development of antivenom is predicated on obtaining sufficient snake venom which poses significant challenges. The venoms of snakes are composed of a mixture of bioactive proteins and peptides that show various biochemical activities. Additionally, snake venom composition varies between different species. Taken together, because of their rich molecular diversity, snake venoms have been a source for drug discovery. Therefore, there is a need to comprehensively understand the complexity of venomous snakes and their association with toxins to aid efficient development of antivenom in order to reduce the clinical risk of snakebites and simultaneously to lay the foundation for the resourceful use of venom. In this thesis, a sequencing-based approach is presented on how to decode the genomic basis of venomous snakes and use the obtained toxin genes for in vitro expression as a proof of concept for the use of recombinant toxin without keeping snakes in captivity or capturing them in the wild. This approach is based on sequencing technology and in vitro expression of recombinant proteins.
V. ammodytes is considered one of the most venomous snakes in Europe, with distribution mainly in Central Asia, the Middle East, Central Europe, and Western Europe. Given the importance of V. ammodytes, we selected V. ammodytes as our research subject. We performed whole genome sequencing and RNA sequencing of V. ammodytes by integrating different sequencing technologies and were able to assemble a chromosome-level genome. From the genome of V. ammodytes, 137 toxin gene homologs from 17 gene families were identified, and 21 of 75 expressed toxin-coding genes specifically expressed in the venom gland were confirmed. Based on the high-quality genome acquired, SVMP toxins with specific structures of V. ammodytes were also validated. In addition, the adaptation of V. ammodytes to the environment has found new perspectives from the genome.

About 45% of the toxin genes identified were not detected to be expressed in snake venom glands of V. ammodytes. Still, all these genes were predicted to have functional domains using sequence analysis. To explore the possible biological activity of these dormant toxin genes, a silent LAAO gene was selected to attempt transgenic in vitro expression using the yeast Pichia Pastoris expression system, thus opening a new path for subsequent resourceful use of (dormant) toxin genes.
It is hoped that the work presented here could help facilitate the development and production of new and effective antivenom and also stimulates innovative thinking to explore the natural resources of venomous snakes.
Original languageEnglish
Place of PublicationKgs. Lyngby, Denmark
PublisherDTU Bioengineering
Number of pages174
Publication statusPublished - 2022

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