10.2 Breakdown of polysaccharide: cellulose

Polysaccharides are polymers of monosaccharides in which the constituent sugars are connected with glycosidic bonds. Because of the number and variety of available sugars and the diversity of bonding possibilities between different carbon atoms of the adjacent sugar residues there is a considerable variety of polysaccharides. There is a matching variety of enzymes, hydrolases or glucosidases, capable of hydrolysing this range of glycosidic links. Enzymes responsible for polymer degradation (any polymer, not just polysaccharide) may employ one of two strategies of attack. They may attack randomly, effectively fragmenting the polymer molecule into a number of oligomers, these are the endo-enzymes, or they may approach terminally, digesting away monomers or dimers, the exo-enzymes.

Cellulose is the most abundant organic compound on Earth and accounts for over 50% of organic carbon; about 1011 tons are synthesised each year. It is an unbranched polymer of glucose in which adjacent sugar molecules are joined by β1→4 linkages (Fig. 1); there may be from a few hundred to a few thousand sugar residues in the polymer molecule, corresponding to molecular masses from about 50,000 to approaching 1 million. Breakdown of cellulose is chemically straightforward, but is complicated by its physical form. Mild acid hydrolysis of cellulose releases soluble sugars, but does not go to completion; oligomers of 100-300 glucose residues remain. The fraction which is readily hydrolysed is called amorphous cellulose while that which is resistant to acid is called crystalline cellulose. Since it influences chemical breakdown, the conformation and three-dimensional structure of cellulose must influence cellulolytic enzyme activity.

Structural formula of cellulose
Fig. 1. Structural formula of cellulose. There may be from a few hundred to a few thousand sugar residues in the polymer molecule, corresponding to molecular masses from about 50,000 to approaching 1 million. Modified from Moore, 1998.

The cellulolytic enzyme (cellulase) complex of white-rot Basidiomycota like Phanerochaete chrysosporium and Ascomycota like Trichoderma reesei consists of a number of hydrolytic enzymes: endoglucanase, exoglucanase and cellobiase (which is a β-glucosidase) which work synergistically and, in both bacteria and fungi, are organised into an extracellular multienzyme complex called a cellulosome (Bégum & Lemaire, 1996). Endoglucanase attacks cellulose at random, producing glucose, cellobiose (a disaccharide made up of two glucose molecules) and some cellotriose (a trisaccharide). Exoglucanase attacks from the non-reducing end of the cellulose molecule, removing glucose units; it may also include a cellobiohydrolase activity which produces cellobiose by attacking the non-reducing end of the polymer. Cellobiase is responsible for hydrolysing cellobiose to glucose. Glucose is, therefore, the readily-metabolised end-product of cellulose breakdown by enzymatic hydrolysis.

The cellulosome, which is highly developed in both bacteria and fungi, is an extracellular molecular machine. In addition to catalytic regions, cellulolytic enzymes contain domains not involved in catalysis, but taking part in substrate binding, multi-enzyme complex formation (so-called ‘docking domains’), or attachment to the cell surface (Shoham et al., 1999). Cellulosomes efficiently degrade crystalline cellulose and associated plant cell wall polysaccharides, provide for attachment to the cell surface and their adhesion to the insoluble substrate provides the individual microbial cell that produced them with a competitive advantage in the utilisation of soluble products. Manipulation of the cellulosome (production of ‘designer cellulosomes’) is seen as a promising way of managing domestic and industrial cellulosic wastes (Bayer et al., 2007).

When grown on cellulose, the white-rot fungi like Phanerochaete chrysosporium produce two cellobiose oxidoreductases; a cellobiose: quinone oxidoreductase (CBQ) and cellobiose oxidase (CBO). Cellobiose oxidase is able to oxidise cellobiose to the δ-lactone, which can then be converted to cellobionic acid and then glucose + gluconic acid; cellobiose δ-lactone can also be formed by the enzyme cellobiose: quinone oxidoreductase. Similar cellobiose-oxidising enzymes, capable of utilising a wide variety of electron acceptors, have been detected in many other fungi, though their role is uncertain. These enzymes are probably of most significance in regulating the level of cellobiose and glucose, the accumulation of which can inhibit endoglucanase activity. The role originally ascribed to CBQ was as a link between cellulose and lignin degradation. Cellobiose oxidase also reduces Fe(III) and together with hydrogen peroxide, generates hydroxyl radicals. These radicals can degrade both lignin and cellulose, indicating that cellobiose oxidase has a central role in degradation of wood by wood-decay fungi.

Brown-rot fungi use a rather different initial cellulolytic system to the hydrolytically-based one employed by the white-rots. Brown-rot fungi are able to depolymerise cellulose rapidly and virtually completely. Even cellulose deep within the walls and protected by lignin polymers is prone to attack. The process seems to depend on hydrogen peroxide (H2O2) secreted by the fungus, and ferrous ions in the wood oxidising sugar molecules in the polymer, thereby fragmenting it and leaving it open to further attack by hydrolytic enzymes. Interestingly, oxalate crystals that coat so many fungal hyphae may be responsible for reducing the ferric ions normally found in wood to ferrous ions, so aiding oxidative cleavage of the cellulose. Although the white-rot fungi produce H2O2 for lignin degradation they do not secrete oxalate and so fail to depolymerise cellulose oxidatively.

Updated December 17, 2016