Page 256 - Advances in Textile Biotechnology

Developments in recombinant silk and other elastic protein fi bers 237

Unfolded α-Helix 3 -Helix β-Sheet β-Turn
10
10.1 Common secondary structural motifs in proteins.

to significant homogeneity in secondary structure (Fig. 10.1) and usually
exhibit important mechanical properties (Altman et al., 2003). By combin-
ing polypeptide sequences derived from these proteins in different ways, a
new protein polymer can be biosynthesized using DNA techniques with
unique physical, mechanical and biological features, such as variations of
folding, chain interactions within the synthetic protein structure, tempera-
ture or pH responsiveness or others (Qiu et al., 2009).
Devising an appropriate strategy at the nucleotide level is essential for

the efficient synthesis of the protein encoding sequence and for producing
a uniform protein product with an optimal quality and yield of the fi nal

protein product. The biosynthesis of any artificial protein generally includes:
(1) constructing a synthetic gene encoding the protein of interest in a
plasmid with tight transcription control; (2) cloning of the recombinant
gene with the necessary transcriptional regulatory elements to competent
cells; (3) screening plasmid containing cells for ones containing the desired
clones and verifying the DNA sequence; (4) transforming the chosen plas-
mids into expression competent host cells; (5) growing appropriate volumes
of host cells and inducing protein expression; (6) purifying the protein of
interest from cell lysates (Mi, 2006).

10.3 Biomimetic design of recombinant proteins

In designing genes encoding repetitive protein-based polymers, the tech-
niques of molecular biology are typically employed to self-ligate monomer
DNA fragments in a process of oligomerization that relies on restriction
enzyme-based approaches. In this instance, the monomer fragments must
be oligomerized in a ‘head-to-tail’ orientation, and can be seamless in
sequence or can contain intervening linkers between the desired repeats.
The approaches to the oligomerization can be classifi ed as:

1 Iterative technique, where a DNA segment is oligomerized in a series
of single, uniform steps, each step growing the oligomer by one unit
length of the monomer gene.

© Woodhead Publishing Limited, 2010