Biodegradation by Microorganisms

Microorganisms capable of lignocellulose degradation are widespread and include mainly fungi, the ascomycetes (e.g., T. reesei), basidiomycetes including white-rot fungi (e.g., P. chrysosporium), brown-rot fungi (e.g., Fomitopsis palustris), and a few anaerobic species (Orpinomyces sp.) that degrade cellulose in GI tracts of ruminants (Yoon et al. 2007; Ljungdahl 2008).

Cellulose Biodegradation

Cellulose degradation is performed by group of glycosyl hydrolases (GHs) that hydrolyzes p-1,4-glycosidic bonds in cellulose called cellulases by retaining and inverting catalytic mechanism. Cellulase belong to GH 12 family (hydrolyze glyco- sidic bonds by the retaining mechanism) and GH 6 family (cellulases use the inverting mechanism) (Bayer et al. 1998; Sandgren et al. 2005). Both catalytic mechanisms include two catalytic carboxylate residues that catalyze the hydrolysis reaction via acid-base catalysis. Cellulases belong mainly to basidiomycetes fungi (ten Have and Teunissen 2001). Table 2.3 includes few examples of cellulases. These fungi produce hydrolytic (for hydrolyzing cellulose and hemicellulose) and oxidative enzymes (for hydrolyzing lignin) and penetrate their mycelia deeply into the ligno- cellulosic substrate. The degradation of lignocellulosic biomass by fungi is either exocellular (in association with the outer cell layer) or extracellular (because of insoluble nature of lignin and carbohydrates). Cellulases are found in aerobic fungi (hyphomycetes, ascomycetes, and basidiomycetes), anaerobic fungi (Bayer et al.

1998), and aerobic and anaerobic bacteria (Gilkes et al. 1991). Filamentous fungi and aerobic bacteria produce cellulases as free molecule called noncomplexed cellulase system, whereas anaerobic bacteria such as C. thermocellum secrete cellulases as complexed system with high molecular weight called cellulosomes (Schwarz 2001). Some of the enzymes derived from these microorganisms can also degrade lignin. Cellulase production by brown-rot fungi such as Poria placenta has been studied in details (Eriksson et al. 1990). No exoglucanases were found in these fungi and endoglucanases have been identified in liquid culture. Depolymerization of cellulose by brown-rot fungi further involves a nonenzymatic oxidative agent such as peroxidases (Charles and Gerald 1989).

Noncomplexed Cellulase System One of the most studied and extensively used noncomplexed cellulase system is the mesophilic fungus Trichoderma reesei (teleo- morph Hypocrea jecorina) which is known as an efficient producer of extracellular enzymes (Bayer et al. 1998). T. reesei includes two cellobiohydrolases (CBH), around seven endoglucanases, and several p-glucosidases. Production of p-gluco- sidase in T. reesei is low, and consequently cellobiose is produced in large amounts which inhibits the activity of endo- and exoglucanases (Gilkes et al. 1991). This end-product inhibition can be reduced when cellobiose is broken down to glucose by adding p-glucosidase from either external sources or by doing fungal co-culturing (Ting et al. 2009). As suggested by Ting et al. (2009), cellulose hydrolysis by T. reesei is performed in three phases:

  • 1. Phase I—chemical and physical modification-change in degree of polymerization, changes in the accessible surface area. Primary enzyme involved in this phase is endoglucanases.
  • 2. Phase II—primary hydrolysis—this phase is slow, involves cellulose degradation to soluble intermediates using cellobiohydrolase (CBH).

Table 2.3 Microorganisms using lignocellulosic carbon sources for lignocellulolytic enzyme production

Microorganism

Substrate

Enzymes produced

Enzyme activities

Reference

FPase

CMCase

P-Glucosidase

Xylanase

T. reesei RUT C30

Roll-milled cotton

Cellulase, xylanase

14 U mL1

150 U mL-1

0.3 UmL-1

NA

Peterson and Nevalainen (2012)

T. reesei QM 9414

Roll-milled cotton

Cellulase, xylanase

10 U mL1

109 U mL1

0.6 U mL1

NA

Peterson and Nevalainen (2012)

T. reesei QM6a

Roll-milled cotton

Cellulase, xylanase

5 U mL1

88 U mL-1

0.3 UmL-1

NA

Peterson and Nevalainen (2012)

T. reesei NG-14

Roll-milled cotton

Cellulase, xylanase

15 U mL1

133 UmL1

0.6 U mL1

NA

Peterson and Nevalainen (2012)

A. niger

Lactose

Xylanase, cellulase

ND

ND

1165 U mL1

212 Ug-1

Gamarra et al. (2010)

A. niger

Lactose

Xylanase, cellulase

ND

ND

1174 UmL-1

217 Ug-1

Gamarra et al. (2010)

A. niger

Sugarcane bagasse

Xylanase, cellulase

Parket al. (2002)

A. fumigatus

Rice straw + wheat bran

Cellulase, xylanase

0.68 IU g-1

14.71 IUg-1

8.51 IUg-1

42.7 IU g-1

Shah and Madamwar (2005)

3. Phase III—secondary hydrolysis—this phase is fast, involves hydrolysis of soluble intermediates produced from phase II to low-molecular-weight oligosaccharides and ultimately to glucose by using p-glucosidase.

Complexed Cellulase System In this category, cellulosomes are produced especially by anaerobic bacteria and have further been found in a few anaerobic fungal spp. such as Neocallimastix, Piromyces, and Orpinomyces (Tatsumi et al. 2006; Watanabe and Tokuda 2010). Cellulosome-producing bacteria belong generally to the phylum Firmicutes and Lachnospiraceae and Clostridiaceae families. Cellu- losomes are protuberances containing enzymes and are produced on the cell wall of the cellulolytic bacteria. A cellulosome consists of two subunits: non-catalytic subunits (scaffoldings) and catalytic or enzymatic subunits. The scaffoldings are termed as functional unit and are reservoir of cohesins that interact selectively with enzymatic subunits. Enzymatic subunits consist of CBD (cellulose binding domains) and CBM (carbohydrates binding modules), responsible for providing binding capacity to the substrate or the cell surface. CBM functions to mediate the binding of the scaffoldings to cellulose substrate (Vandamme 2001). Recognition domains, cohesin and dockerin, are called as receptor/adaptor pairs and are complementary to each other. The interaction between cohesin and dockerin forms the positional selfassembly of the cellulosome; they are specific for each bacterial species (Lynd et al. 2002; Arai et al. 2006). Third system consists of two cellulolytic bacteria, anaerobic bacteria such as Fibrobacter succinogenes, present in rumen, and aerobic bacteria Cytophaga hutchinsonii present in soil. They contain endocellulase and lack CBMs (Xie et al. 2007; Wilson 2008).

 
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