30
quantities of heterologous β- glucosidase, generally sourced from Aspergillus niger
(Knauf and Moniruzzaman 2004; Rana et al. 2014). It is often difficult to gauge the
optimal combinations of the different component enzymes that give maximal hydrolysis of a given feedstock and hence the highest sugar concentrations. Mostly, the
optimizations of enzyme cocktails have been empirical, with different mixtures
being analyzed for their efficacy in hydrolysis of the target feedstock. This often
presents the issue of having to try several combinations of component enzymes
experimentally, before arriving at the optimal one. Recently, high throughput methods have been used for such optimizations and the US Great Lakes Bioenergy
Research Center platform called the GENPLAT (Great Lakes Bioenergy Research
Center Enzyme Platform) is an approach aimed at addressing this issue. GENPLAT
allows rapid assays and optimization of enzyme mixture tailored for different pretreatments, feedstock, and combinations of them (Banerjee et al. 2010).
Cost reduction of commercial enzymes for biomass hydrolysis often has to
employ a combination of different strategies that cover use of cost-effective substrates/carbon source, efficient fermentation strategy that minimizes catabolite
repression, use of cheaper but efficient inducer(s),enzymes with improved activities
and/or stability, enzyme modification to reduce nonspecific binding, development
of cocktails with higher efficiency, additives that enhance the enzyme activity,
enzyme reuse, and onsite production (Fig. 1.8).
As has been stated previously, the cost reduction of biomass-hydrolyzing
enzymes is not just about reducing the production cost of cellulases, but more economy can be achieved through multiple steps that improve the hydrolysis step and
through usage of lesser enzyme per unit biomass to be hydrolyzed (Marcuschamer
et al. 2012; Liu et al. 2016). Nevertheless, a major contribution can definitely come
from reduction of enzyme production cost. Cheaper fermentation strategy like
solid-state fermentation may definitely improve the cost of production, but if the
enzyme needs elaborate downstream processing, this may not be cost effective.
Most of the biomass-hydrolyzing enzyme preparations are not used in their purified
form and the only downstream processing steps would be filtration and concentration, since accessory activities are equally important as the main cellulase and hemicellulase activities and crude preparations often perform better for hydrolysis of
pretreated biomass. Direct use of the moldy bran for biomass hydrolysis was already
described under the section for solid-state fermentation (Zhuang et al. 2007,
Singhania et al. 2015). The major advantage of solid-state fermentation can be the
use of cost-effective substrate/inducers in the form of native or pretreated lignocellulosic biomass (Cunha et al. 2012). Cellulases being inducible enzymes, appropriate inducers are added in the fermentation medium for production of enzymes. In
most of the industrial production scenarios using T. reesei, the common inducer
added is lactose, which happens to be the only economically viable soluble inducer.
It should be noted that the best inducer for T. reesei is sophorose, which is very
expensive for any commercial-scale production. However, this issue could be
resolved by generating the inducer the way nature does it: by transglycosylation of
sugars mediated by beta glucosidases. The patent from Danisco describes exactly
that, and large quantities of inducer (sophorose) were demonstrated to be generated
R.K. Sukumaran et al.
quantities of heterologous β- glucosidase, generally sourced from Aspergillus niger
(Knauf and Moniruzzaman 2004; Rana et al. 2014). It is often difficult to gauge the
optimal combinations of the different component enzymes that give maximal hydrolysis of a given feedstock and hence the highest sugar concentrations. Mostly, the
optimizations of enzyme cocktails have been empirical, with different mixtures
being analyzed for their efficacy in hydrolysis of the target feedstock. This often
presents the issue of having to try several combinations of component enzymes
experimentally, before arriving at the optimal one. Recently, high throughput methods have been used for such optimizations and the US Great Lakes Bioenergy
Research Center platform called the GENPLAT (Great Lakes Bioenergy Research
Center Enzyme Platform) is an approach aimed at addressing this issue. GENPLAT
allows rapid assays and optimization of enzyme mixture tailored for different pretreatments, feedstock, and combinations of them (Banerjee et al. 2010).
Cost reduction of commercial enzymes for biomass hydrolysis often has to
employ a combination of different strategies that cover use of cost-effective substrates/carbon source, efficient fermentation strategy that minimizes catabolite
repression, use of cheaper but efficient inducer(s),enzymes with improved activities
and/or stability, enzyme modification to reduce nonspecific binding, development
of cocktails with higher efficiency, additives that enhance the enzyme activity,
enzyme reuse, and onsite production (Fig. 1.8).
As has been stated previously, the cost reduction of biomass-hydrolyzing
enzymes is not just about reducing the production cost of cellulases, but more economy can be achieved through multiple steps that improve the hydrolysis step and
through usage of lesser enzyme per unit biomass to be hydrolyzed (Marcuschamer
et al. 2012; Liu et al. 2016). Nevertheless, a major contribution can definitely come
from reduction of enzyme production cost. Cheaper fermentation strategy like
solid-state fermentation may definitely improve the cost of production, but if the
enzyme needs elaborate downstream processing, this may not be cost effective.
Most of the biomass-hydrolyzing enzyme preparations are not used in their purified
form and the only downstream processing steps would be filtration and concentration, since accessory activities are equally important as the main cellulase and hemicellulase activities and crude preparations often perform better for hydrolysis of
pretreated biomass. Direct use of the moldy bran for biomass hydrolysis was already
described under the section for solid-state fermentation (Zhuang et al. 2007,
Singhania et al. 2015). The major advantage of solid-state fermentation can be the
use of cost-effective substrate/inducers in the form of native or pretreated lignocellulosic biomass (Cunha et al. 2012). Cellulases being inducible enzymes, appropriate inducers are added in the fermentation medium for production of enzymes. In
most of the industrial production scenarios using T. reesei, the common inducer
added is lactose, which happens to be the only economically viable soluble inducer.
It should be noted that the best inducer for T. reesei is sophorose, which is very
expensive for any commercial-scale production. However, this issue could be
resolved by generating the inducer the way nature does it: by transglycosylation of
sugars mediated by beta glucosidases. The patent from Danisco describes exactly
that, and large quantities of inducer (sophorose) were demonstrated to be generated
R.K. Sukumaran et al.