Surface Modification of Cellulose Acetate with Cutinase and ...

module (CBM) of Cellobiohydrolase I, or the bacterial CBM of Endoglucanase C. The
new recombinant cutinase fused to the fungal CBM presented higher hydrolysis of
cellulose diacetate and improved the colour levels of the treated fabrics (Subchapter
2.5).
The Chapter 3 describes the design of enzymatic-based technologies for wool fibres
finishing industrial applications. Wool has the intrinsic characteristic to felt and shrink
due to its scaly structure; the chlorine-Hercosett is the commercial process used to
modify the scales of wool fibres with the purpose of providing resistance to felting and
shrinkage. There have been several attempts to replace this chlorine process by
proteases, in order to degrade scales, providing wool with anti-felting and antishrinkage
characteristics. However, proteases commercially available can diffuse inside
the fibre causing unacceptable damages. In this thesis two novel approaches were
followed to increase molecular weight of the subtilisin E by genetic engineering. Polyenzymes
composed of two and four subtilisin E coding sequences fused in frame were
constructed. Additionally, another chimeric subtilisin was obtained by 3´-terminus
fusion with the nucleotidic sequence coding for the human neckdomain of surfactant
protein D. All chimeric subtilisins were cloned and overexpressed into E. coli but the
soluble and active forms were not attained, regardless the expression system or the
strain used, under the culture conditions tested (Subchapter 3.2). Subchapter 3.3
describes the fusion of subtilisin E gene in frame with the DNA coding an elastin-like
polymer containing 220 repeats of the monomer VPAVG. With this strategy the
construction of a chimeric enzyme presenting a molecular weight above 116 kDa was
achieved. Wool yarns treated either with commercial or chimeric enzyme showed a
size-dependent diffusion process: the commercial enzyme penetrated into wool cortex
while the chimeric one was retained at the surface, in the cuticle layer. These results
represent a major achievment: the production of a recombinant high molecular weight
protease, for wool surface controlled-hydrolysis, is reported for the first time.
Chapter 4 presents a general discussion, the major conclusions and gives some
perspectives for continuing the work in this research field.
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