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In situ Reversible Aggregation of Extracellular Cellobiase in the Filamentous Fungus Termitomyces clypeatus

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Title In situ Reversible Aggregation of Extracellular Cellobiase in the
Filamentous Fungus Termitomyces clypeatus
 
Creator Banik, Samudra Prosad
Pal, Swagata
Ghorai, Shakuntala
Chowdhury, Sudeshna
Majumder, Rajib
Mukherjee, Soumya
Khowala, Suman
 
Subject Drug Development/Diagnostics & Biotechnology
 
Description Cellobiase (E.C. 3.2.1.21), is a widely exploited
industrial glycosidase with a major role in biofuel industry.
Its stability and shelf life are major bottlenecks in
achieving a superior formulation for industry. In the
filamentous fungus Termitomyces clypeatus, the enzyme is
secreted in a co-aggregated form with sucrase; the
separation of this co-aggregation results in substantial loss
of the enzyme’s activity. The aim of the present study was
to examine the mode of aggregation of the secreted
cellobiase-sucrase coaggregate and its role in the
stabilization of cellobiase. Transmission electron microscopy
and dynamic light scattering of purified co-aggregates
revealed reversible, concentration driven self-aggregation
of the extracellular enzymes to form larger entities.
However, the intracellular enzyme aggregates were rigid,
non-interacting, and possessed a higher percentage of
disulphide bonds. Circular dichroic spectra of the two coaggregates
indicated no significant difference in secondary
structures. Self-association increased the stability of
extracellular aggregates towards heat by 1.5 fold, SDS by
4 ~ 7 fold, and chaotropic agents, by 1.5 ~ 2 fold, than the
intracellular counterpart. The Km of extracellular aggregate
varied between 0.29 and 0.45 mM as a result of
spontaneous aggregation and disaggregation, whereas that
of intracellular aggregate was 0.22 mM irrespective of its
concentration status. In situ detection of cellobiase in
native PAGE revealed two activity bands of the
extracellular enzyme, which indicated a minimum of two
active dissociated aggregate species, as compared to a
single band for the intracellular enzyme. These studies are
believed to improve the understanding of aggregation of
the fungal glycosidases, which remains to be a blackbox, to
increase the efficacy of these enzymes
 
Date 2012
 
Type Article
PeerReviewed
 
Format application/pdf
 
Identifier http://www.eprints.iicb.res.in/1715/1/BIOTECHNOLOGY_AND_BIOPROCESS_ENGINEERING__17___(5)_925%2D936;2012[28].pdf
Banik, Samudra Prosad and Pal, Swagata and Ghorai, Shakuntala and Chowdhury, Sudeshna and Majumder, Rajib and Mukherjee, Soumya and Khowala, Suman (2012) In situ Reversible Aggregation of Extracellular Cellobiase in the Filamentous Fungus Termitomyces clypeatus. Biotechnology and Bioprocess Engineering, 17 (5). pp. 925-936.
 
Relation http://dx.doi.org/10.1007/s12257-012-0002-9
http://www.eprints.iicb.res.in/1715/