Knottins provide useful scaffolds or leads for drug design

  • ‣ Knottins are exceptional in that they are very small proteins yet with particularly well-defined scaffolds and remarkably high stability.
  • ‣ Also remarkable is the fact that knottins with very similar 3D structures have virtually no sequence identity except for cysteines. This observation has led to the conclusion that the knottin scaffold is very sequence tolerant.
  • ‣ These remarkable features suggest that knottins can provide excellent lead molecules or elementary scaffold in drug design studies [Chiche et al., 2004; Craik et al., 2006; Werle et al., 2006; Moore et al., 2012], and in biotechnological applications [Cox et al., 2016; Moore et al., 2013; Glotzbach et al., 2013; Barba et al., 2012].
  • ‣ Main efforts along this way are outlined below.
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Cell internalization

Circular permutations

Computer simulations

Homology modeling



Protein engineering

Circular permutations

Linearization of the circular peptide Kalata B1

Cyclic peptides can be linearized by breaking of a peptide bond, usually in solvent accessible loops. Structure and stability of the linear peptide give informations on the importance of the loops in the folding and the stability of the circular peptide. Such linearizations are similar in spirit to the circular permutations performed on non-cyclic peptides (see below).
Six different linear analogs of the cyclic peptide Kalata B1 have been synthesized [Daly & Craik, 2000]. Four linear peptides are able to fold correctly indicating that the broken loops are not essential for folding. It is shown that the two linear peptides that do not fold correspond to peptides in which the disulfide macrocycle typical of the Knottins has been disrupted. This highlight the importance of the knot in Knottins.

Circular permutation of EETI-II

A similar approach has been applied to the non-cyclic EETI-II Knottin [Chiche, Heitz & Strub, unpublished results]. In such cases the native termini of the peptide are linked by a regular peptide bond (usually with addition of a short peptide link), and new termini are introduced somewhere else in the peptide resulting in true circular permutations. Here the C- and N-termini are connected with addition of three glycines and the 17-18 peptide bond has been cleaved. The resulting permuted peptide has been shown to fold correctly, indicating that the 16-19 β-turn is not essential for folding.