Abstract
Animal venoms are complex mixtures containing peptides and proteins known as toxins, which are responsible for the deleterious effect of envenomations. Across the animal Kingdom, toxin diversity is enormous, and the ability to understand the biochemical mechanisms governing toxicity is not only relevant for the development of better envenomation therapies, but also for exploiting toxin bioactivities for therapeutic or biotechnological purposes. Most of toxinology research has relied on obtaining the toxins from crude venoms; however, some toxins are difficult to obtain because the venomous animal is endangered, does not thrive in captivity, produces only a small amount of venom, is difficult to milk, or only produces low amounts of the toxin of interest. Heterologous expression of toxins enables the production of sufficient amounts to unlock the biotechnological potential of these bioactive proteins. Moreover, heterologous expression ensures homogeneity, avoids cross-contamination with other venom components, and circumvents the use of crude venom. Heterologous expression is also not only restricted to natural toxins, but allows for the design of toxins with special properties or can take advantage of the increasing amount of transcriptomics and genomics data, enabling the expression of dormant toxin genes. The main challenge when producing toxins is obtaining properly folded proteins with a correct disulfide pattern that ensures the activity of the toxin of interest. This review presents the strategies that can be used to express toxins in bacteria, yeast, insect cells, or mammalian cells, as well as synthetic approaches that do not involve cells, such as cell-free biosynthesis and peptide synthesis. This is accompanied by an overview of the main advantages and drawbacks of these different systems for producing toxins, as well as a discussion of the biosafety considerations that need to be made when working with highly bioactive proteins.
Highlights
Animal venoms present a treasure trove of biologically active compounds that have evolved to perform highly specialized biochemical tasks, in the contexts of defense against predators and prey capture (Arbuckle et al, 2017; Rivera-de-Torre et al, 2020)
We present the possibilities offered by the principal heterologous expression systems for the heterologous expression of toxins as well as strategies for producing toxins without cells, such as cell-free biosynthesis or chemical synthesis of peptides
This strategy has been successfully applied to the expression of α-latrotoxin, a 130 kDa neurotoxin produced by widow spiders that is extremely difficult to extract from the venom gland in large amounts (Volynski et al, 1999)
Summary
Animal venoms present a treasure trove of biologically active compounds that have evolved to perform highly specialized biochemical tasks, in the contexts of defense against predators and prey capture (Arbuckle et al, 2017; Rivera-de-Torre et al, 2020) These venoms are complex mixtures of peptides and proteins displaying toxic activity, commonly known as toxins, salts, and small metabolites, such as neurotransmitters and nucleosides. Given the challenges of obtaining rare and low-abundance toxins, other approaches must be taken for procuring animal toxins to fully exploit the potential that lies within their diversity In this relation, heterologous expression of toxin genes in a laboratory setting presents an exciting and promising alternative to extracting animal toxins from their natural source. We highlight some of the most promising research efforts involving toxin expression, e.g., antivenom research, development of bioinsecticides, toxinderived drug development, and the bioethical considerations surrounding such research activities
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