19
anaerobic cellulosome-bearing microorganism Clostridium thermocellum (Berger
et al. 2007).
1.3.3 Other Systems for Cellulose Hydrolysis
Certain microorganisms use strategies that do not fit into the complexed or noncomplexed cellulose hydrolysis paradigms. Typical examples are the certain members of the genus Caldicellulosiruptor and the genus Fibrobacter, each of which
possess a different strategy for cellulose hydrolysis other than the classical complexed or free enzyme paradigms. In Caldicellulosiruptor bescii, the cellulolytic
machinery is midway between cellulosomes and free cellulases in that it consists of
secreted cellulases that are multimodular, containing multiple binding and catalytic
domains. The catalytic domains can be with different activities (Brunecky et al.
2013). The dominant cellulase in this organism – CelA – is a complex thermostable
enzyme containing N terminal glycosyl hydrolase family 9 (GH9) endoglucanase
domain, 3 GH3 CBMs, and a terminal C terminal GH48 catalytic domain, the latter
an exoglucanase. The family 9 and 48 catalytic domains are highly synergistic as
described in Clostridium thermocellum (Vazana et al. 2010). The multifunctional
combination of enzyme activities is functionally distinct from cellulosomes and the
Caldicellulosiruptor enzymes exist as free enzymes not associated with the cell
(Young et al. 2014). CelA is now known to act both by conventional cellulase processivity and excavation of cavities into the surface of the biomass substrate
(Brunecky et al. 2013).
Even more intriguing is the entirely different cellulolytic mechanism employed
by the rumen bacterium Fibrobacter succinogenes. The organism contains genes for
endoglucanases, but does not have genes for exoglucanases or processive endoglucanases, both of which are needed for release of cellobiose from cellulose chains
(Ransom Jones et al. 2012). Interestingly, the anaerobic cellulose degrader does not
possess a cellulosome; nor does it have CBMs on the enzymes it produces (Burnet
et al. 2015). It is now known that certain outer membrane proteins in F. succinogenes called the fibro-slime proteins are involved (Jun et al. 2007), and these proteins along with type IV pilin structures mediate the attachment of the organism to
the substrate (cellulose). Cellulose chains are broken by a protein complex present
on the outer cell wall, which includes catalytic function, and the individual cellulose
chains released through this are transported into the perisplasmic space via an ABC
transporter. Once here, they are depolymerized by endoglucanases, eliminating the
need for processive enzymes or exoglucanases (Wilson 2009). While a lot more is
yet to be learned about the mechanism of cellulose degradation in Fibrobacter, there
seems to be a consensus that this is one of the most efficient mechanisms for cellulose degradation among microbes and holds great promise for use in biomass conversion strategies. The different mechanisms employed by microorganisms are
represented in Fig. 1.6.
1 Enzymes for Bioenergy
anaerobic cellulosome-bearing microorganism Clostridium thermocellum (Berger
et al. 2007).
1.3.3 Other Systems for Cellulose Hydrolysis
Certain microorganisms use strategies that do not fit into the complexed or noncomplexed cellulose hydrolysis paradigms. Typical examples are the certain members of the genus Caldicellulosiruptor and the genus Fibrobacter, each of which
possess a different strategy for cellulose hydrolysis other than the classical complexed or free enzyme paradigms. In Caldicellulosiruptor bescii, the cellulolytic
machinery is midway between cellulosomes and free cellulases in that it consists of
secreted cellulases that are multimodular, containing multiple binding and catalytic
domains. The catalytic domains can be with different activities (Brunecky et al.
2013). The dominant cellulase in this organism – CelA – is a complex thermostable
enzyme containing N terminal glycosyl hydrolase family 9 (GH9) endoglucanase
domain, 3 GH3 CBMs, and a terminal C terminal GH48 catalytic domain, the latter
an exoglucanase. The family 9 and 48 catalytic domains are highly synergistic as
described in Clostridium thermocellum (Vazana et al. 2010). The multifunctional
combination of enzyme activities is functionally distinct from cellulosomes and the
Caldicellulosiruptor enzymes exist as free enzymes not associated with the cell
(Young et al. 2014). CelA is now known to act both by conventional cellulase processivity and excavation of cavities into the surface of the biomass substrate
(Brunecky et al. 2013).
Even more intriguing is the entirely different cellulolytic mechanism employed
by the rumen bacterium Fibrobacter succinogenes. The organism contains genes for
endoglucanases, but does not have genes for exoglucanases or processive endoglucanases, both of which are needed for release of cellobiose from cellulose chains
(Ransom Jones et al. 2012). Interestingly, the anaerobic cellulose degrader does not
possess a cellulosome; nor does it have CBMs on the enzymes it produces (Burnet
et al. 2015). It is now known that certain outer membrane proteins in F. succinogenes called the fibro-slime proteins are involved (Jun et al. 2007), and these proteins along with type IV pilin structures mediate the attachment of the organism to
the substrate (cellulose). Cellulose chains are broken by a protein complex present
on the outer cell wall, which includes catalytic function, and the individual cellulose
chains released through this are transported into the perisplasmic space via an ABC
transporter. Once here, they are depolymerized by endoglucanases, eliminating the
need for processive enzymes or exoglucanases (Wilson 2009). While a lot more is
yet to be learned about the mechanism of cellulose degradation in Fibrobacter, there
seems to be a consensus that this is one of the most efficient mechanisms for cellulose degradation among microbes and holds great promise for use in biomass conversion strategies. The different mechanisms employed by microorganisms are
represented in Fig. 1.6.
1 Enzymes for Bioenergy