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Review

Cellulases from psychrophilic microorganisms: a review

Ramesh C. Kasana and Arvind Gulati

Institute of Himalayan Bioresource Technology, Council of Scientific and Industrial Research,
Palampur (HP), India

Cellulases are hydrolytic enzymes that catalyze total hydrolysis of cellulose into sugars.
Cellulases are produced by various groups of microorganisms and animals; however, psychro-
philes are the ideal candidates for the production of enzymes active at low temperature and
stable under alkaline conditions, in the presence of oxidants and detergents, which are in large
demand as laundry additives. The cellulases from psychrophiles also find application in
environmental bioremediation, food industry and molecular biology. Research work on
cellulase has been done over the last six decades, but there is no exclusive review available on
the cellulases from psychrophiles. This review is an attempt to fill this gap by providing all the
relevant information exclusively for cellulases from psychrophiles, with a focus on the present
status of knowledge on their activity, molecular characteristics, gene cloning, statistical expe-
rimental designs, crystal structure, and strategies for the improvement of psychrophilic cel-
lulases.

Keywords: Cellulase / Detergents / Characterization / Crystal structure / Improvement / Psychrophiles

Received: September 24, 2010; accepted: November 20, 2010

DOI 10.1002/jobm.201000385

Introduction

*

The major part of the earth’s surface provides conditions
that are extreme due to one or the other environmental
factor. These extreme environments, which are detri-
mental to higher life forms, are generally inhabited by
microorganisms able to grow and survive under these
harsh conditions. Low temperature is very common
among the extreme environments, both natural and
man-made. These low-temperature environments are
inhabited by the cold-adapted microorganisms known
as psychrophiles. In the past couple of decades, it has
been recognized that cold-adapted microorganisms pro-
vide a broad biotechnological potential, offering nu-
merous economic and ecological advantages over the
use of those organisms and their respective enzymes
that operate at higher temperatures [25, 28].
Cellulose, which is one of the most abundant carbo-
hydrates produced by plants, provides the major re-
newable energy source on the planet Earth. Its turnover,

Correspondence: Ramesh Chand Kasana, Institute of Himalayan Bio-
resource Technology (CSIR), Palampur-176061, India
E-mail: [e-mail], [e-mail]
Fax: [phone]

therefore, plays an important role in the global carbon
cycle for all living organisms. Cellulose synthesis and
recycling in the cold environments account for a large
proportion of the carbon cycle because approximately
80% of the biosphere and more than 90% of the marine
environments have temperatures lower than 5 °C. In
natural environments, the degradation of cellulose is
mainly carried out by cellulases produced by various
bacteria and fungi [11–13, 38]. Natural sources of cellu-
lases generally contain at least three groups of cellu-
lolytic activities, including endoglucanase, β-1,4-cello-
biohydrolase, and β-glucosidase. The cold-active enzy-
mes are becoming more attractive compared to their
mesophilic counterparts because of their potential in-
dustrial applications, and they represent the lower
natural limits of protein stability. These enzymes are
useful tools for studies in the field of protein folding
[9]. A large number of cold-active enzymes have been
isolated from the psychrophiles, but the cold-active
cellulases are short of study up to now.

Psychrophiles with cellulase activity
Organisms growing at low temperature have been
known for a long time. The term psychrophile was first

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used by Schmidt-Nielson for microorganisms that are
able to grow and multiply at 0 °C [35]. According to
Morita [29], psychrophiles are microorganisms that
show optimal growth at or below 15 °C, maximal
growth at or below 20 °C, and minimal growth at or
below 0 °C. However, the microorganisms that grow at
or below 5 °C but show optimal growth at or above
20 °C have been defined as psychrotrophs. Since psy-
chrotrophs are facultative psychrophiles, the term psy-
chrophiles in this review includes both true and facul-
tative psychrophiles. The psychrophilic microorganisms
are not restricted to cold environments, but are distrib-
uted over almost all kinds of environments. Cellulase-
producing psychrotrophic microorganisms have been
isolated from various environments.
In 1996, the first report on a low-temperature-active
cellulase produced by the fungus Acremonium alcalophi-
lum of soil origin had appeared [16], followed by the
isolation and purification of endo-β-glucanase from the
psychrophilic yeast Rhodotorula glutinis of soil origin [32].
Subsequently, cellulase-producing psychrophiles be-
longing to different bacterial and fungal genera have
been isolated and characterized. The various psychro-
trophic microorganisms producing cellulase are listed
in Table 1.
Microbial enzymes are often more useful than en-
zymes from plants and animals because of the great
variety of catalytic activities available, higher yields,
possible ease of genetic manipulation, regular supply

Table 1. Cellulase-producing psychrophiles.

Isolate identification

Source

Reference

Acremonium alcalophilum

Soil [16]

Arthrobacter sp.

Signy Island

[5]

Cadophora malorum, Cladosporium

cladosporioides, Cladosporium sp.

Wood and swab [7]

Chryseomonas luteola

Celeriac (Apium

graveolens)

[26]

Clostridium sp.

Cattle manure

digester

[3]

Coprinus psychromorbidus

[22]

Fibrobacter succinogenes Rumen [24]

Flavobacterium sp.

Soil sample

[34]

Geomyces sp.

Wood and swab [7]

Paenibacillus sp.

Mud samples

[36]

Pedobacter sp.

Dirt sample

[34]

Penicillium chrysogenum

Yellow Sea

[17]

Penicillium expansum, Penicillium

roquefortii, Penicillium sp.

Wood and swab [7]

Pseudoalteromonas haloplanktis

[44]

Pseudoalteromonas sp.

Mud [48]

Pseudoalteromonas sp.

Sea sediment

[49]

Rhodotorula glutinis

Soil [32]

Shewanella sp. G5

Munida

subrrugosa

[6]

due to the absence of seasonal fluctuations, and the
rapid growth of microorganisms on economical media.
Microbial enzymes are also more stable than their cor-
responding plant and animal enzymes, and their pro-
duction is relatively more convenient and safer [47].
Cold-adapted microbial strains have been isolated most-
ly from the antarctic and polar regions, which repre-
sent permanently cold environments. Other potential
sources of cold-active cellulases are the mud and deep-
sea sediment microorganisms. A Clostridium strain iso-
lated from a cold-adapted manure biogas digester was
able to grow at temperatures as low as 5 °C and as high
as 50 °C [3], whereas the CMCase produced by the fun-
gus Acremonium alcalophilum was active even at 0 °C [16].
We have isolated cellulose-producing psychrotrophic
bacteria belonging to the genera Paenibacillus and Pseu-
domonas from the cold environments in the Western
Himalaya (Fig. 1).

Figure 1. Cellulase production by bacteria belonging to the genera
Paenibacillus and Pseudomonas.

Basic properties of cellulases from psychrotrophs
In the last one and a half decade, psychrotrophic mi-
croorganisms belonging to various groups have been
screened for cellulase production, purification, and char-
acterization. But the information is scanty on the puri-
fication and characterization of cellulases from psy-
chrotrophs. The first report on the characterization of a
low-temperature-active cellulase appeared in year 1996
for Acremonium alcalophilum [16]. The cellulase of Acre-
monium alcalophilum showed maximal activity at 40 °C
and pH 7.0, the enzyme was cold active, retaining more
than 20% activity even at 0 °C [16]. Subsequently cellu-
lases have been isolated and characterized from differ-
ent psychrophilic microorganisms. The optimum tem-
perature for their activity generally ranges from 20
to 40 °C (Table 2), with one study reporting a higher

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optimum temperature for activity (50 °C) by a cellu-
lase from the psychrotrophic yeast Rhodotorula glutinis
[32]. As is clear from Table 2, most of the cellulases
produced by psychrophilic microorganisms showed
optimum activity in the acidic to neutral pH range
(pH 4.5–7.0), with the exception of cellulases from
Paenibacillus sp., Pseudoalteromonas sp. MB-1, and Shewa-
nella sp., which showed their optimum activity in the
neutral to alkaline pH range [6, 36, 48]. Cellulases from
psychrophilic microorganisms reported up to now have
a size range of 40–93 kDa (Table 2). The effect of vari-
ous metals, activators, and inhibitors on the cellulase
activity is given in Table 2. The endoglucanase (Cel9P)
from Paenibacillus showed maximal activity at pH 6.5 and
a temperature of 35 °C for the degradation of CMC-Na
and was stable in buffers of pH 5–10 (at 4 °C for 1 h),
with little loss of activity under alkaline conditions;
moreover, it was adapted to low temperature, retaining
around 65% of its maximal activity at 5 °C [10].
As is clear from the above-reviewed literature, most
of the cellulases from psychrophiles are cold active,
showing optimum activity at temperatures around 35–
40 °C. The cellulases included in this review are pro-
duced extracellularly, and all these cellalose-producing
microorganisms are not true psychrophiles but faculta-
tive psychrophiles and can grow at 30–35 °C. Psychro-
phily is an evolutionary development from thermophily.
Thus, the extracellular enzyme activity may not be
the highest but sufficient to support growth at the

temperature optimum for bacterial growth. The cellu-
lases are active in nature at acidic to slightly alkaline
pH ranging from 4.5 to 8.0. The cellulosome-like en-
zyme from Eisenia foetida is a multienzyme complex of
CMCase with β-glucosidase, β-1,3-glucanase, and β-xylo-
sidase, with a molecular mass of 150 kDa on gel filtra-
tion under non-reducing condition. The CMCase activ-
ity in the purified enzyme complex at 15 °C was 44% of
the activity obtained at the optimum temperature, and
the optimum pH was 5.0 [41].
Cellulases have been incorporated into detergents since
the early 1990s for laundry purposes. During textile
washing, cellulases remove cellulose microfibrils form-
ed during manufacturing and washing of the cotton-
based cloth [23]. Cellulases can also be used as color-
brightening and softening agents. Considering the
present market scenario in the detergent industry, in-
corporating cellulases with proteases in detergent for-
mulations would be advantageous for enhancing the
washing performance of detergents. The detergent in-
dustry is now focusing on cellulases for lower (cooler)
washing temperatures (30–40 °C) and reduction in wa-
ter consumption [37]. Hence, for using the cellulases as
detergent additives, the enzyme should be active under
low temperature, stable under alkaline conditions, and
compatible with oxidants and detergents. If the cellulase
is resistant to degradation (hydrolysis) by proteases, that
would be of added advantage for detergent formulation
and reduce the overall cost of detergent formulation.

Table 2. Kinetic properties of cold-active cellulases from psychrophiles.

Psychrophilic

microorganism

Cellulolytic

enzyme

Temp. opti-

mum (°C)

pH

optimum

Molecular

weight (kDa)

Activators/

inducers

Inhibitors Reference

Acremonium

alcalophilum

CMCase 40 7.0

–

Glucose –

[16]

Arthrobacter sp.

β

-Glucosidase 35

–

93

[5]

Clostridium sp.

Endoglucanase,

β

-glucosidase,

filter paper

cellulase

20 5–6

– –

–

[3]

Fibrobacter

succinogenes

Endoglucanase 25

5.5

55

–

–

[24]

Paenibacillus sp.

strain C7

β

-Glucosidase 30–35 7–8 –

–

–

[36]

Paenibacillus sp.

BME-14

Endoglucanase 35

6.5

60

Ca

2+

, Mg

2+

, Pb

2+

,

Mn

2+

, dithiothrei-

tol, β-mercapto-

ethanol

Hg

2+

, Cu

2+

, EDTA [10]

Pseudoalteromonas

sp. MB-1

Endoglucanase

35 7.2

– –

–

[48]

Pseudoalteromonas

sp. DY3

Endoglucanase 40

6–7

52

–

–

[49]

Rhodotorula glutinis Endoglucanase 50

4.5

40

Fe

3+

, Mn

2+

Hg

2+

, Al

3+

, Ca

2+

,

Fe

2+

, Pb

2+

, EDTA,

EGTA

[32]

Shewanella sp. G5

β

-Glucosidase 37

8.0

–

–

–

[6]

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Cloning and expression of the cellulase genes
Gene cloning is a rapidly progressing technology that
has been instrumental in improving our understanding
of the structure-function relationship of genetic sys-
tems. It offers an excellent technique for the exploita-
tion and regulation of genes. Many of the industrially
important enzymes are now produced from genetically
engineered microorganisms. For the commercial utili-
zation of enzymes, the achievement of higher enzyme
yields is required at low capital cost. Gene cloning and
overexpression can play a significant role in reducing
the production cost of enzymes. Generally, cellulases
are composed of three domains: the cellulose-binding
domain and the catalytic binding domain separated by
a linker region [40]. Garsoux and coworkers cloned,
sequenced and expressed in Escherichia coli the cold-
adapted cellulase CelG from Pseudoalteromonas haloplank-
tis and reported that this cellulase was composed of
three distinct regions: an N-terminal catalytic domain
belonging to the glycosidase family 5, a C-terminal
cellulose-binding domain belonging to the carbohydra-
te-binding module family 5, and a linker of 107 resi-
dues, one among the longest linkers found in cellu-
lases, resulting in increased substrate accessibility to
the catalytic domain by increasing the surface of cellu-
lose available to a bound enzyme molecule [11]. More-
over, the long linker region has been found to play a
key role in adaptation to cold by the psychrophilic cel-
lulase from Pseudoalteromonas haloplanktis [39]. The celX
gene encoding the cold-active cellulase (endoglucanase)
isolated from psychrotrophic Pseudoalteromonas sp. pro-
duced a protein consisting of 492 amino acids [49]. The
gene celA of Pseudoalteromonas sp. encoding a cold-adapt-
ed endogluanase was cloned and expressed in E. coli
BL21 [48]. The psychrophilic cellulase CelX differed from
its mesophilic counterpart in having a longer linker
sequence containing 73 additional residues. Hou and
coworkers isolated the cellobiohydrolase 1 gene (cbh1)
from the cold-adapted fungus Penicillium chrysogenum
and cloned and expressed it in Saccharomyces cerevisiae,
but the recombinant enzyme accumulated intracellu-
larly and was not secreted into the medium [17]. Gener-
ally, extracellular enzyme production is favored over
intracellular enzyme production, the latter adding to
the production cost of the process.

Optimization for cellulase production through
statistical experimental designs
For making use of cellulases from psychrotrophs for
the industry, it is of immense importance to optimize
the parameters for cellulase production. As far as cellu-
lases from psychrotroph are concerned, there is not

even a single publication exclusively focusing on the
optimization of cellulase production from psychro-
trophs. In one of the papers on cellulase optimization,
among the various carbon sources tested,

D

-glucose

showed maximum protease production by Rhodotorula
glutinis while beef extract and yeast extract as nitrogen
sources were found to increase the cellulase produc-
tion. Maximum enzyme production was observed at
pH 5 after incubation at 20 °C for 96 h [32]. Generally,
the medium optimization is carried out by using the
classical method by changing one independent variable
at a time and keeping the other factors constant, lead-
ing to a considerable increase in enzyme yields. But this
approach is burdensome and time consuming; in addi-
tion, it ignores the interaction of various physico-
chemical parameters and hence it does not depict the
complete effects of these parameters on the process. On
the other hand, the statistical approach using response
surface methodology for process optimization serves
the purpose by finding the optimal conditions in any
given system by a set of independent variables and
establishing the relationship between more than one
variable. An overall 1.5-fold increase in cellulase pro-
duction was reported from Trichoderma harzianum by
employing a two-level fractional factorial design (FFD)
and using six factors, i.e. substrate and co-substrate
concentration, temperature, initial pH, inoculum size,
and agitation rate [4]. Optimization of the medium for
the production of cellulase by the mutant Trichoderma
reesei WX-112 using response surface methodology re-
sulted in an increase in cellulase activity from 7.2 to
10.6 IU/ml [15].
A model developed and validated using response
surface methodology for the optimization of cellulase
production was found reliable for maximizing the cel-
lulase production by Penicillium waskmanii F10-2 [14].
Response surface methodology has also been employed
for optimization of other enzymes produced by various
microorganisms, including the optimization of cold-
active protease production by psychrophilic Colwellia
sp., with the enzyme production coinciding with the
predicted value [45]. So for the optimization of cellu-
lase production from psychrophiles, the statistical ap-
proach using response surface methodology should be
exploited in the future.

Insight into the crystal structure of cold-adapted
cellulases
One of the major goals of a structural study is to clarify
the molecular basis of adaptation to low temperatures.
The essential structural features of psychrophilic cellu-
lases that contribute to their high psychrophilic activ-

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ity might have developed through an evolutionary pro-
cess that led to the separation of a group of psychro-
philic strains from their relatives. Cold-adapted cellu-
lases probably are structurally modified by increasing
the flexibility of the polypeptide chain, enabling an
easier accommodation of substrates at low tempera-
ture. The fundamental issues concerning the molecular
basis of cold activity and the interplay between flexibil-
ity and catalytic efficiency are of importance in the
study of structure-function relationships in enzymes.
Such issues are often approached through comparison
with the mesophilic or thermophilic counterparts, by
site-directed mutagenesis and three-dimensional (3D)
crystal structure analysis [31, 33, 46]. Therefore, 3D
structural comparisons between mesophilic and psy-
chrophilic cellulases are required in order to under-
stand how proteins can adapt to low temperatures. The
first report on the structure of a psychrophilic protein,
a cold-active α-amylase from the antarctic psychrophile
Alteromonas haloplanctis A23, came in the year 1996 [2],
which was followed by a preliminary crystal structure
determination of the alkaline protease from the antarc-
tic psychrophile Pseudomonas aeruginosa [42]. However,
the 3D structure of psychrophilic cellulases was not
known till 2003, when a first paper appeared on the
crystallization and preliminary X-ray crystallographic
studies of a psychrophilic cellulase from Pseudoaltero-
monas haloplanktis [44]. The X-ray diffraction data at
1.8 Å resolution for the catalytic core domain showed
the space group to be P 2

1

2

1

2

1

, with refined unit-cell

parameters a = 135.1, b = 78.4, c = 44.1 Å. Violot et al.
[43] studied the structural properties of the full-length
cellulase from P. haloplanktis by using both X-ray diffrac-
tion and the small-angle X-ray scattering method. The
results of small-angle X-ray scattering showed that the
linker region between the catalytic binding module and
the cellulose-binding module is unstructured and un-
usually long and flexible compared to its mesophilic
counterpart, indicating that the linker plays a signifi-
cant role in the cold adaptation of this psychrophilic
cellulase. The longer linker in the cellulase from the
psychrophilic P. haloplanktis as compared to the cellu-
lase from Erwinia chrysanthemi is a highly flexible struc-
ture due to various factors, like negatively charged
residues, the absence of positively charged residues,
and only a small number of hydrophobic residues [39,
43]. Due to the highly flexible structure of the linker,
the catalytic module and the cellulose-binding domain
are free to move to a larger extent, which helps in their
correct positioning with respect to the cellulose sub-
strate at low temperature below 25 °C [43]. The struc-
ture-based alignment of the catalytic modules of the

cellulases from P. haloplanktis (Cel5GCM) and Erwinia
chrysanthemi (Cel5GCM) showed that Cel5GCM has two
two-residue insertions (T64-S65 and W274-N275) com-
pared to Cel5ACM [43]. Due to insertion of these residue
pairs in each loop, β-turns are formed that are unique
to Cel5GCM.
The comparison of the crystal structures of cold-
adapted enzymes and their homologous enzymes from
mesophilic and/or thermophilic organisms is yielding
fruits to elucidate the structural basis of temperature
adaptation. Structural comparisons support the original
hypothesis that cold-adapted proteins appear to contain
fewer or weaker intramolecular interactions, which is
assumed to facilitate global or local flexibilities in the
protein molecules. The most frequently reported struc-
tural differences involve interactions such as fewer
intra- or inter-subunit salt bridges, less compact hydro-
phobic packing in the protein core, longer surface loops
and fewer prolines in such loops, an increased number
of glycine clusters, a reduced number of proline/ar-
ginines, improved solvent interactions through addi-
tional surface (especially negative) charges, increased
solvent exposure of apolar surfaces, and a better acces-
sibility of the active site. As the crystal structures of
cellulases are being solved, this will considerably facili-
tate the design of rational engineering strategies for
enzymes with desired properties (enzymes with altered
specificity, stability, biophysical properties).

Commercially available cold-active cellulases
The companies producing cold-active cellulases to-
gether with the cellulases’ characteristics and uses are
listed in Table 3. Dyadic’s ROCKSOFT

TM

Antarctic and

Antarctic LTC cellulases are two cold-water cellulases
for the stonewashing process to take place at 40 °C or
below and hence resulting in energy savings for the
textiles manufacturers, versus the traditional stone-
wash cellulases that typically are run at higher tem-
peratures [19]. The cold-water neutral cellulase UTA-90
and the neutral cellulase UTA-88 produced from a gene-
tically engineered microbial strain by Hunan Youtell
Biochemical, Ltd., are low-backstaining products func-
tioning at 30–40 °C over a broad pH range, with a shelf
life of 12 months at temperatures below 25 °C [21]. The
Retrocell RecoP cold-temperature powdered and buf-
fered cellulase is an engineered cellulase product that
has been specially designed to wash denims at low
temperatures and in a wide range of pH, which results
in very low backstaining and good contrast. This pow-
dered enzyme system contains a pH buffer for main-
taining near neutral pH (5.5–6.0) during the wash
cycle. Also, Retrocell ZircoN, a cold-temperature liquid

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Table 3. Properties of commercially available cold-active cellulases.

Enzyme name

Temp. optimum (°C) pH optimum Uses

Commercial producer

ROCKSOFT

TM

Antarctic LTC conc.

40

5.5

Softening cotton fabrics, woven and

knits. Increases the “stonewash”

effect on rotary-washed garments

using a significant amount of pumice

stones and/or other abrasives.

Dyadic International, Inc.

Cold-water neutral

cellulases UTA-88 &

UTA-90

30–40

6.5–7.8

“Stonewash” of jeans, offering high

contrast texture patterns with low

backstaining.

Hunan Youtell Biochemical

Co. Ltd.

Retrocell RecoP &

Retrocell ZircoN

35–45

6.0–6.5

Used at low temperatures to retain

the original indigo color of denim

and backstaining is maintained at

the minimum level.

Used in denim “stonewashing” with

or without stones and additives to

produce the desired abrasion levels.

EpyGen Biotech

Siltex 30443

Ultrazyme CLD

30–60 (range)

5.5–7.5

Used for cold-water wash to obtain

good color pull, high abrasion, low

fabric strength loss, and low back-

staining.

Paradise Industrial

Corporation

cellulase, is an engineered cellulase capable of washing
indigo denims at low temperatures (30 °C or lower) and
in a wide range of pH, with good contrast and low
backstaining [20]. Siltex 30443 Ultrazyme CLD is a cellu-
lase enzyme for cold-water stonewashing, working in
the temperature range of 30–60 °C and at pH 5.5–7.5
[18].

Future perspectives
Day by day, the market demand for industrial enzymes
is increasing, necessitating the screening of various
sources for enzymes with different functionalities.
There are different strategies that can be employed to
obtain efficient enzymes with the desired properties for
industrial applications. (i) The huge unexplored biodi-
versity is an invaluable resource for biotechnological
innovations, which can play a significant role in the
search for new strains of microorganisms required in
the industry. The exploitation of the biodiversity to pro-
vide microorganisms that produce enzymes well suited
for various applications would always remain a valu-
able option, with emphasis on isolating/culturing bac-
teria expressing cellulase activity from unexplored
niches, by employing various techniques. Metagenomic
libraries should be established from the cold-environ-
ment sites rich in cellulosic materials, for obtaining
novel cellulases from the uncultured microorganisms.
(ii) Strain improvement by either conventional mutage-
nesis or through recombinant DNA technology is an-
other option. Penicillium janthinellum NCIM 1171 was sub-
jected to mutation, and five selected mutants showed
an approximately twofold increase in activity of both
FPase and CMCase in shake flask cultures [1], and the

strain improvement of Acremonium cellulolyticus for cellu-
lase production by mutagenesis resulted in higher en-
zyme yields with higher filter paperase (FPase) activities
(17.8 U/ml) compared to the parent strain C-1 (12.3
U/ml) [8].
Recombinant DNA technology was used to improve
the thermal stability of the Clostridium cellulovorans cel-
lulosomal endoglucanase (EngB) in vitro by recombina-
tion with non-cellulosomal endoglucanase EngD [30].
(iii) Another approach is protein engineering, where
site-directed mutagenesis of the cellulase gene can be
employed to improve the yield, stability, and the cata-
lytic properties of the enzyme. The hyperthermostable
endoglucanase Cel5A from Thermotoga maritima sub-
jected to site-directed mutagenesis and carbohydrate-
binding module (CBM) engineering showed a shift in
optimal pH from 5 to 5.4 in five single mutants, with
one mutant displaying 10% higher activity than the
native strain. After domain engineering by binding two
CBMs, one from Trichoderma reesei and the other from
Clostridium stercorarium, to Cel5A, the CBM-engineered
Cel5A showed 14–18-fold higher hydrolytic activity to-
wards Avicel [27]. Hence, these approaches should be
followed for targeting cold-active cellulases with activi-
ties and stabilities over a broad range of pH values and
temperatures.
The use of protein engineering requires thorough
knowledge of the protein structure for site-directed
mutagenesis to be carried out. However, for the major-
ity of cellulases in general and cellulases from psychro-
trophs, we do not have structural information available
and our knowledge about predicting protein structure
and function is still in its infancy. Therefore, random

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© 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

www.jbm-journal.com

mutagenesis/directed evolution will continue to be a
leading technique to modify cellulases for some more
time in future.

Acknowledgements

The authors are grateful to the Director of the Institute
of Himalayan Bioresource Technology, Palampur, for
support and encouragement. The Council of Scientific
and Industrial Research is also acknowledged for finan-
cial support under the CSIR Network Project “Exploita-
tion of India’s Rich Microbial Diversity” (NWP 006).

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((Funded by

• Council of Scientific and Industrial Research, CSIR
Network Project “Exploitation of India’s Rich Microbial
Diversity” (NWP 006)))