Production, p urification and characterization of cellulase from Aspergillus species
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Date
2017
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
DEPARTMENT OF BIOCHEMISTRY AND BIOCHEMICAL ENGINEERING, JACOB SCHOOL OF BIOTECHNOLOGY AND BIOENGINEERING, SAM HIGGINBOTTOM UNIVERSITY OF AGRICULTURE, TECHNOLOGY AND SCIENCES, ALLAHABAD - 211007
Abstract
Optimization parameters (pH, tem
perature
, moisture content, incubation period) and effect
of co
-
inducers on production of cellulase (FPase, CMCase, and cellobiose
) produced by
A. flavus
and
A. japonicum
isol
ated from soils of Allahabad, U.
P.
via
solid state fermentation
(SSF) using
sugarcane bagasse, wheat straw and paper waste as substrates
was studied. In
crude enzyme extracts
both strains exhibited higher endogl
ucanase (CMCase) activity as
compared to cellobiase and total cellulase activity (FPase). The specific activities of the three
enzymes also followed a similar pattern
i.e.
highest in paper waste as compared
to
the
s
ugarcane baggase and wheat straw
. The
hig
hest production of cellulase was obtained with
A.
flavus
when 5.0g paper waste in 1:3 basal medium ratio growth medium (30mL); 1.0% (w/v)
yeast extract and cellulose as co
-
inducers was used at 30°C, pH 4.5, 96 hrs of SSF
CMCase
1150.81 ± 6.26 IU/L cellobia
se 670.44±5.8IU/L and FPase 598.53
±5.56 IU/L)
.
The
optimized conditions were used to extract and purify
CMCase by
a three step procedure,
the
purification factor was 462.5
after gel filtration yield 4%
Molecular
weight of CMCase was
found
to be ~
23 KDa
as
determined by SDS
-
PAGE
. The purified endoglucanase
retained
90%of its maximum activity after 4
5min of incubation at 50°
C. The endoglucanase activity
stimulated
66%
by
Co
2+
however,
Hg
2+
competitively inhibited CMCase with a K
i
of 2μM.
The
value of K
m
and V
max
were
increased
16.66%
by
33.33%
after
purification.
The
efficiency of crude cellulase produced by
A
. flavus
and
A. japonicum
in bioconversion was
also compared.
With purified
enzyme the
degree of saccharification was
lower than crude
enzyme for all the
substrates. Though, paper waste was saccharified the most
among all
substrates
. Molecular characterization of the two
Aspegillus species
revealed two
novel
strains and were assigned
N
CBI accession no. KF716164
(A. flavus
) and
KF716165
(
A.
japonicum
)
.
Key
words:
Bioconversion,
CMCase, Cellobiase, FPase
,
SSF, Saccharification
.
Description
DOCTOR OF PHILOSOPHY
IN
BIOCHEMISTRY
Keywords
null