Page 36 - Advances in Textile Biotechnology

Design and engineering of novel enzymes for textile applications 15

Other workers have followed the same strategy of construction of fusion
proteins using CBMs from different sources. The application of CBMs in
several areas of biotechnology has increased recently because CBMs are
independently folded domains and a priori they can function perfectly when
fused to other proteins. Moreover, their ligands are abundant and renew-
able materials with good properties, and they are normally low-priced
(Shoseyov and Warren, 1997). One of the most important areas is biopro-

cessing, because large-scale purification and recovery of biological mole-
cules continues to be a challenge. CBMs can be expressed as an affi nity tag

for protein immobilization, processing and purification using cellulose as a

matrix in many formats, from affinity chromatography to two-phase liquid
separations (McCormick and Berg, 1997). Targeting of compounds to poly-
saccharides that are present in many daily products, in particular cellulose,
is also an important application of CBMs. This area of application includes,
among others, denim stonewashing and the targeting of enzymes that do

not possess a natural affinity to cotton in laundry detergents, (Berry et al.,
2001; Cavaco-Paulo, 1995; Kalum and Andersen, 2000; von der Osten et al.,
2000a,b).
Degani and collaborators (2004) developed a new method to measure

the scouring efficiency of cotton based on the fusion of a cellulose-binding
domain with the β-glucuronidase protein (CBD-GUS). The chimeric
enzyme was used as an effective tool for measuring scouring and wettabil-
ity of cotton fabrics. CBD-GUS binding increased with hydrophilicity of
the fabric owing to the increasing harshness of the scouring treatment.
This quantitative method combines both simplicity and sensitivity in con-
trast to qualitatively inaccurate techniques or those requiring expensive
instrumentation.
Matamá et al. (2010) presented a novel and promising approach to
increase cutinase adsorption on cellulose acetate fibers by fusing cutinase

with two distinct (bacterial or fungal) CBM. They found an increase in
reactivity and hydrophilicity of fibers treated with chimeric enzyme com-

pared with the native cutinase.
The application of CBMs is far from being exhausted and will be further
expanded as the understanding of these binding domains increases.

1.4.4 Proteases: subtilisins
Subtilisins are a family of alkaline serine proteases secreted by a variety of
Bacillus species (Siezen and Leunissen, 1997). These enzymes catalyze the
hydrolysis of peptide and ester bonds through formation of an acyl-enzyme
intermediate.
The objective of cloning bacterial protease genes was mainly the over-
production of enzymes for the detergent and leather industries and, more

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