A Spider Toxin Exemplifies the Promises and Pitfalls of Cell-Free Protein Production for Venom Biodiscovery

Arthropod venoms offer a promising resource for the discovery of novel bioactive peptides and proteins, but the limited size of most species translates into minuscule venom yields. Bioactivity studies based on traditional fractionation are therefore challenging, so alternative strategies are needed....

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Bibliographic Details
Published in:Toxins 2021-08, Vol.13 (8), p.575
Main Authors: Lüddecke, Tim, Paas, Anne, Talmann, Lea, Kirchhoff, Kim N, von Reumont, Björn M, Billion, André, Timm, Thomas, Lochnit, Günter, Vilcinskas, Andreas
Format: Article
Language:English
Subjects:
Arthropods
Binding sites
Biological activity
biotechnology
Biotechnology - methods
cell-free expression
Cell-Free System - physiology
E coli
Fractionation
Gel electrophoresis
Invertebrates
knottin
Mass spectrometry
Mass spectroscopy
neglected venomous arthropods
New technology
Peptides
Protein Biosynthesis - physiology
Protein expression
Protein synthesis
Proteins
Scientific imaging
Sodium lauryl sulfate
spider venom
Spider Venoms - chemistry
Spiders
Synthesis
Synthetic biology
Synthetic Biology - methods
Toxins
Venom
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Summary:Arthropod venoms offer a promising resource for the discovery of novel bioactive peptides and proteins, but the limited size of most species translates into minuscule venom yields. Bioactivity studies based on traditional fractionation are therefore challenging, so alternative strategies are needed. Cell-free synthesis based on synthetic gene fragments is one of the most promising emerging technologies, theoretically allowing the rapid, laboratory-scale production of specific venom components, but this approach has yet to be applied in venom biodiscovery. Here, we tested the ability of three commercially available cell-free protein expression systems to produce venom components from small arthropods, using U -sicaritoxin-Sdo1a from the six-eyed sand spider as a case study. We found that only one of the systems was able to produce an active product in low amounts, as demonstrated by SDS-PAGE, mass spectrometry, and bioactivity screening on murine neuroblasts. We discuss our findings in relation to the promises and limitations of cell-free synthesis for venom biodiscovery programs in smaller invertebrates.
ISSN:2072-6651
2072-6651
DOI:10.3390/toxins13080575