Page 64 - Advances in bioenergy (2016)

which is called the dockerin module involved in enzyme interaction with the scaffolding
proteins. Scaffoldins are large multidomain, multifunctional proteins that recruit catalytic
proteins interacting with GH dockerin domains and improve complex affinity for the substrate
and catalytic efficiency via CBMs. Such a complex architecture enables the enzymatic
components to act in a synergistic and coordinated manner via intra- and intermolecular
interactions and makes the cellulosomes the most efficient biochemical systems for cellulose
degradation.

Oxidative Enzymes for Cellulose Degradation

Recently, a revolutionary mode of action was discovered for the hydrolysis of cellulose,
revealing the mechanism that the members of the known GH61 family follow (Table 2.1), using
oxidative chemistry. This chemistry occurs in the cleavage of chitin (a fibrous, crystalline
polymer of β-1,4-N-acetylglucosamine residues) by the bacterial chitin-binding protein 21
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(CBP21) that belongs to CBM family 33. This novel mode of action opens up the crystalline
polysaccharide material that is inaccessible for hydrolysis by other GHs. A homologue to
CBP21 in Streptomyces coelicolor was shown to act synergistically with cellulases in the
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digestion of cellulose. The CBP21 oxidative mechanism led to the assumption that the
structurally similar GH61 enzymes, which were known for many years as weak
endoglucanases, might also follow the same mode of action on crystalline cellulose. The first
example that supported this assumption, reported that some members of GH61 family could
significantly enhance the effectiveness of common cellulases in the breakdown of complex
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lignocellulosic substrates. Interestingly, another study revealed that the combination of a
Thermoascus aurantiacus GH61 with a Humicola insolens cellobiose dehydrogenase resulted
in cellulose cleavage, yielding a mixture that included reducing end-oxidized and nonreducing
20
end-modified cellooligosaccharides. In addition, the authors showed a synergistic effect of
both Thielavia terrestis GH61 and cellobiose dehydrogenase on microcrystalline cellulose
20
hydrolysis by T. terrestris canonical cellulases. It was hypothesized that an oxidoreductive
cellulolytic system coexists with the well-studied fungal cellulases resulting in efficient
19
lignocellulose conversion. In GH61 enzymes, copper is the most likely redox center and
taking in consideration that the catalytic metal-binding site is conserved, it is highly probable
that members of both the GH61 and CBP21 families are oxidative enzymes that target
recalcitrant plant cell wall material such as cellulose (Figure 2.2). It has been demonstrated
that the presence of lignin, which presents a significant source of reductant residues, is
probably responsible for the increase in activity of GH61 from Myceliophthora thermophila
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by acting as electron donors. This boosting effect was first described by the use of the
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reductant ascorbic acid by increasing the efficiency of CBP21. These fungal derived, copper-
depended oxidative enzymes lacking hydrolase activity, are recently named polysaccharide
monooxygenases (PMO) that catalyze the O -dependent oxidative cleavage of recalcitrant
2
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polysaccharides. In addition to this surprising discovery in the theory of cellulose hydrolysis,
a new type of enzymes with possibly a very important role in cellulose utilization was
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discovered in the M. thermophila supernatant termed aldonolactonases. These recently
discovered enzymes catalyze the hydrolysis of glucono-δ-lactone and cellobiono-δ-lactone that