Chemical synthesis and recombinant protein expression


  • ‣ Since knottins are considered as interesting leads or scaffolds in drug design, it is necessary that these miniproteins can be obtained easily
  • ‣ As they are very small proteins, many knottins were obtained by chemical synthesis
  • ‣ Several linear and cyclic knottins were also obtained using recombinant technologies
  • ‣ Main techniques used to prepare knottins (including cyclic knottins) are outlined below

Knottins can be obtained either:


Synthesis

Expression

Protease inhibitors

Cyclotides

Toxins

Others

Toxins

Recombinant toxin production

Few toxins have been produced as recombinant proteins. Conotoxin MVIIA, which is used as an analgesic (Ziconotide, commercial name PRIALT) has been expressed in Escherichia coli as a fusion protein with thioredoxin [Zhan et al, 2003].
Conotoxin It7a from Conus litteratus has been expressed and purified in Escherichia coli from a fusion protein with thioredoxin and His-tag [Pi et al, 2007].

Early synthesis of ω-conotoxin GVIA

As conotoxins are usually small peptides and because many bear post-translational modifications, solid-phase chemical synthesis is the most used method to produce conotoxin samples. Early syntheses appeared in the late 80s [Rivier et al, 1987].