Cellulase (Types, Sources, Mode of Action & Applications)

Cellulase (Types, Sources, Mode of Action & Applications)
Cellulase is a class of enzyme that catalyzes the cellulolysis i.e., hydrolysis of cellulose. Celulase
is a multiple enzyme system consisting of endo – 1, 4 –β–D – glucanases and exo – 1, 4 –β– D –
glucanases along with cellobiase (β– D – glucosideglucano hydrolase).
Types of Cellulases
On the basis of fractionation studies on culture filtrate have demonstrated that, there are ‘three’
major types of enzymes involved in the hydrolysis of native cellulose to glucose, namely: Others
are produced by the some animals and plants.
1. Endo-glucanase
Endo-glucanase (endo –β– 1, 4 – glucanase, endo –β– 1, 4 – D – glucan – 4 – glucanohydrolase,
E.C.3.2.1.4) ofen called CMC ase, hydrolysis carboxyl methyl cellulose or swollen cellulose, due
to which there is a rapid decrease in chain length along with a slow increase in reducing groups.
Endoglucanse also acts on cellodextrins, the intermediate products of cellulose hydrolysis and
converts them to cellobiase and glucose.
2. Exo-glucanase
Exoglucanase (Exo–β– 1, 4 – D – glucan – 4 – cellobiohydrolase, E.C. 3.2.1.91) degrades
cellulose by splitting off the cellobiost units from the nonreducing end of the chain.
Cellobiohydrolase does not degrade cotton rapidly, but can effect considerable saccharification
of micro crystalline celluloses.
3. β – glucosidase / Cellobiase
β– glucosidase ( E.C.3.2.1.21) completes the process of cellulose hydrolysis by cleaving
cellobiose and removing glucose from the non - reducing ends of oligosaccharides. Complete
degradation of cellulose to glucose requires the synergistic action of all the three components.

4. Animal Cellulases
It is now clear that some cellulolytic insects produce their own cellulases and also harbor
symbiotic cellulolytic microorganisms. Cellulolytic animals play an important role in plant cell
wall degradation by grinding up the biomass material into small particles, thus making it much
easier for the cell wall-degrading enzymes in their digestive system to access and hydrolyze the
sugar polymers. In effect, the animals are pretreating the biomass they ingest, making it more
accessible to their enzymes and to those of their symbiotic microorganisms. It is not exactly
known how the cellulases produced by the different organisms present in insect guts function in
cellulose degradation. A recent paper described sequencing the metagenome from the hindgut of
a termite, which showed that many different plant cell wall-degrading enzymes from many
different microorganisms were present. Some cellulolytic termites (Macrotermitinae) utilize
aerobic symbiotic cellulolytic fungi (Termitomyces) to break down plant material in their nests
and then eat the fungi and residual plant material. There also are a few other animals that
produce cellulases, including several types of mollusks particularly snails, slugs, and shipworms.
5. Plant Cellulases
Plants produce cellulases, and different plant cellulases can have quite different roles in plant
physiology; some plant cellulases clearly function in degrading cellulose, such as those involved
in fruit ripening or leaf abscission in the fall. Others are involved in growth and remodeling of
plant cell walls and their exact role in these processes is not clear. Finally, there is a membrane-
bound cellulase, which is required for cellulose biosynthesis. Its exact role in this process is not
known, although it might release cellulose chains from cellulose synthetase. Most plant
cellulases do not contain a carbohydrate-binding module (CBM); however some of those that
clearly function in cellulose degradation do contain one or more CBMs.

Chemical and Physical Mutagens Used for Strain Improvement
There are three main methods of strain improvement available to increase the extracellular
production of cellulases, i.e., (1) mutagenesis and selection, (2) genome shuf- fling, and (3) gene
cloning.
Table: Methods for strain improvement
Strains producing Cellulases
Table 1: Various sources of Cellulases
Microbe Strains
Fungi Soft rot fungi
Aspergillus niger; A. nidulans; A. oryzae; A. terreus; Fusarium solani; F.
oxysporum; Humicola insolens; H. grisea; Melanocarpus albomyces;
Penicillium brasilianum; P. occitanis; P. decumbans; Trichoderma reesei; T.
longibrachiatum; T. harzianum; Chaetomium cellulyticum; C. thermophilum;
Neurospora crassa; P. fumigosum; Thermoascus aurantiacus; Mucor

circinelloides; P. janthinellum; Paecilomyces inflatus; P. echinulatum;
Trichoderma atroviride
Brown rot fungi
Coniophora puteana; Lanzites trabeum; Poria placenta; Tyromyces palustris;
Fomitopsis sp. White rot fungi Phanerochaete chrysosporium; Sporotrichum
thermophile; Trametes versicolor; Agaricus arvensis; Pleurotus ostreatus;
Phlebia gigantean
Bacteria Aerobic bacteria
Acinetobacter junii; A. amitratus; Acidothermus cellulolyticus; Anoxybacillus
sp.; Bacillus subtilis; B. pumilus; B. amyloliquefaciens; B. licheniformis; B.
circulan; B. flexus; Bacteriodes sp.; Cellulomonas biazotea; Cellvibrio gilvus;
Eubacterium cellulosolvens; Geobacillus sp.; Microbispora bispora;
Paenibacillus curdlanolyticus; Pseudomonas cellulosa; Salinivibrio sp.;
Rhodothermus marinus
Anaerobic bacteria
Acetivibrio cellulolyticus; Butyrivibrio fibrisolvens; Clostridium thermocellum;
C. cellulolyticum; C. acetobutylium; C. papyrosolvens; Fibrobacter
succinogenes; Ruminococcus albus
Actinomycetes Cellulomonas fimi; C. bioazotea; C. uda; Streptomyces drozdowiczii; S.
lividans; Thermomonospora fusca; T. curvata
Mode of Action
There appear to be at least three different ways by which cellulolytic microorganisms degrade
cellulose.
 Most aerobic cellulolytic microorganisms use the free cellulase mechanism in which they
secrete a set of individual cellulases (six to ten), each of which contains a CBM joined by a
flexible linker peptide to the CD.

 Most anaerobic microorganisms use the cellulosomal mechanism producing large (>1 million
MW) multienzyme complexes, called cellulosomes, which are usually bound to the outer
surface of the microorganism.
 The third strategy appears to be used by at least two cellulolytic bacteria: Cytophaga
hutchinsonii, an aerobe and Fibrobacter succinogenes, an anaerobe. The DOE Joint Genome
Institute has determined the DNA sequence of the C. hutchinsonii genome
(http://genome.jgi-psf.org). C. hutchinsonii codes for a number of cellulase genes, most of
which do not encode a CBM, none of which encodes a dockerin domain, and all of which
appear to code for endoglucanases.
Mechanism of cellulose hydrolysis by microorganisms Cellulases are enzymes which able to
break down cellulose by hydrolyse b-1-4 glycosidic bonds of cellulose polymer. The complete
hydrolysis of cellulose into glucose requires the synergetic action of at least three enzymes,
endoglucanases preferably, attack amorphous regions and randomly cleave the internal bonds of
the glycan chains, thus providing reducing or nonreducing ends of cellooligosaccharides for
cellobiohydrolases to attack. CBH then hydrolyses those chain ends in the processive manner,
yielding cellobiose as the major product. Lastly, b-glucosidase further hydrolyses cellobiose to
glucose and also releases glucose from the nonreducing ends of soluble cellooligosaccharides.

Figure 1: Mode of action of microbial Cellulases
A

There is a high degree of synergy between cellobiohydrolases (exoglucanases) and
endoglucanases, which is required for the efficient hydrolysis of cellulos. The products of
endoglucanases and cellobiohydrolases, which are cellodextrans and cellobiose, respectively, are
inhibitory to the enzyme’s activity. Thus, efficient cellulose hydrolysis requires the presence of
bglucosidases which cleaves the final glycosidic bonds producing glucose (end product)
Typically, cellobiose and cellodextrins are taken up by the microorganism and internally cleaved
via cellodextrin phosphorylases or cellobiose phosphorylases to create glucose monophosphate,
which is energetically favoured. Some bacteria also produce intra- or extracellular b-glucosidases
to cleave cellobiose and cellodextrins and produce glucose to be taken up or assimilated by the
cell. Mechanistically, the reactions catalysed by all cellulases are suggested to involve general
acid–base catalysis by a carboxylate pair at the enzyme active site, though different in structure.
One residue acts as a general acid and protonates the oxygen of the o-glycosidic bond; at the
same time, the other residue acts as a nucleophile. Depending on the distance between the two
carboxylic groups, either inverting
(
10 A˚ distances) or retaining
(
5 A˚ -distances) mechanisms are observed in cellulases. Moreover, the involvement of multiple
enzymes with a wide range of substrate specificities enables constant enzymatic actions on
lignocellulosics Unlike soluble substrates that can diffuse the active sites of enzymes, cellulose is
insoluble; thus, cellulases, on the contrary, have to diffuse, attach, and move the segment of the
cellulose polymer to their active sites. Most cellulases are modular proteins comprising discrete

catalytic modules that typically appended one or more carbohydrate-binding modules (CBMs)
joined by a flexible linker. The CBM functions as a cellulose probe, in which the main
responsibility is binding the enzyme to the cellulose and increasing the effective concentration of
enzymes on the surface of the cellulose. In addition, some CBMs are known to possess the
ability to disrupt crystalline cellulose. Therefore, the presence of CBMs appears to be important
in enhancing the enzymatic activity towards insoluble polysaccharides, as well as crystalline
cellulose.
Application of Cellulases
Microbial Cellulases find applications in various industries as shown in Figure 1. However the
detail applications of Cellulases are mentioned in Table 1.

Figure 1: Biotechnological applications of Cellulases (Behera et al., 2017)
Application of
Cellulases
Food Industry
Animal Feed
Industry
Textile &
Laundry
industry
Beer & wine
industry
Pulp & paper
Industry
Agriculture
Industry
R&D
Industry
Bio fuel
Industry
Pharmaceutic
al Industry
Waste
management
Industry
rDNA
Technology

Table 1: Application of Cellulases in various industries (Khud et al., 2011)
Global Market of Cellulases
Cellulases are currently the third largest industrial enzyme worldwide, by dollar volume, because
of their use in cotton processing, in paper recycling, as detergent enzymes, in juice extraction,
and as animal feed additives. However, cellulases will become the largest volume industrial
enzyme if ethanol from lignocellulosic biomass through the enzymatic route becomes a major
transportation fuel. The demand for cellulases is consistently on the rise because of its diverse
applications.

Table 2: Companies producing cellulases on commercial scale

Cellulase (Types, Sources, Mode of Action & Applications)

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Cellulase (Types, Sources, Mode of Action & Applications)