De Novo Design of Proteins - Peptide and Protein Design for Biopharmaceutical Applications - page 228

geometry of the peptide-attachment points, or that the template merely
controls the oligomeric state of the assembly (overall topology), for
example that four and five helical strands on the template would give
4-helix vs. 5-helix bundles, respectively.
While this template strategy for protein design has led to very fasci-
nating science, it is notable that no X-ray crystal or NMR structures of
TASPs have been reported so far, which has limited the development of
this field. The lack of NMR structures could in part be due to the often
degenerated structures (i.e. multiple copies of the same peptide
sequence) in these proteins. However, very recently, small-angle X-
ray scattering (SAXS) has provided novel information on the topology
of so-called carboproteins, vide infra . Although these are 'low-resolu-
tion' structures that do not provide atomic resolution, they do provide
key information on the overall fold of the proteins, and hence their
topology.

Table 6.3 Some amphiphilic peptide sequences used in template-based
proteins

Entry Sequence Reference

1 LEALEKALKEALAKLG Mutter [119]

2 LKALKEAFEKAMAELG Mutter

3 EELLKKLEELLKKG Sherman [131]

4 -CEKLLKELKELLEKG-NH 2 Sherman [130]

CGGGEELLKKXEELLKKG 1

5 Sherman [130]

6 GEELLKKLEELLKKGGGC Sherman

7 Ac-EALEKALKEALAKLGG-H Carboprotein [140]

8 Ac-YEELLKKLEELLKKAG-H Carboprotein [145]

1 X: Leu, Ile, Nle or Val; anchoring through Cys

The underlying concept in TASPs is the preorganization of
peptide strands on a molecular scaffold, to reduce the entropy of the
construct's unfolded state, thus to reduce the entropic cost in going from
an unassembled conformation to a constrained protein structure [118].
The templates originally utilized by Mutter and coworkers [118-
120] were short, linear peptides inspired by gramicidin S with two
antiparallel b-sheet segments, which were eventually cyclized to pro-
vide cyclo [-Pro-Gly-Lys-Ala-Lys-Pro-Gly-Lys-Ala-Lys-] and cyclo [- D -
Pro-Gly-Lys-Ala-Lys- D -Pro-Gly-Lys-Ala-Lys-], and their derivatives.
Turn mimetics have also been used in these types of template. The