296
J. Querellou et al.
screening procedures and directed evolution have been applied with success by several research groups and SMEs as well as larger corporations (Diversa, Genencor,
Degussa, Henkel, etc.) to different targets, enzymes and/or novel natural products.
Table 8.3 gives an overview of the most significant metagenomic contributions
during the recent years.
Though limited to ocean surface microbes, the Sorcerer II/GOS expedition represented a milestone for metagenomics and the team predicted more than six million
proteins in the GOS data, approximately twice the number of proteins present at
that time in the databases (Yooseph et al. 2007). The practical utility of the Sorcerer
II results for marine biotechnology is hard to estimate and their exploitation for
that purpose will take many more years to come to fruition. One of the primary
goals of the Sorcerer II studies was to understand the rate of discoveries of protein
families with the increasing number of protein predictions. From the relationship
observed, they concluded that their sampling of the protein universe is far away
from saturation, meaning that many more protein families remain to be discovered.
This conclusion is subject to controversy and Koonin (Koonin 2007) suggested comparing not only the increase in the number of protein clusters with the number of
sequences observed in the Sorcerer II data but also adding the rate of increase of
orthologs. The virome (viral fraction of the microbiome in a given habitat) of several marine regions has been studied and this has also revealed a completely novel
diversity of proteins (Breitbart et al. 2002, 2004, Angly et al. 2006). The results from
genomics projects dedicated to Archaea viral proteins with unknown functions have
shown that marine viruses represent a reservoir of interest for the discovery of new
folds and novel structures and therefore novel protein families (Vestergaard et al.
2008).
With the advent of metagenomic approaches, the microbial diversity is increasingly accessible and we might expect an exponential growth in the number
of biocatalysts potentially useful for the industry, as already demonstrated by
some recent work on nitrilases, lipases and esterases (Robertson et al. 2004,
Bertram et al. 2008), at least when functional screens are efficient. Note, however, that in contrast to global marine metagenome surveys, these results were
obtained in direct collaboration with industry, with clearly defined biotechnological
goals.
There are two main obstacles to the biotechnological exploitation of marine
extremophiles. Screening methods (sequence-based and activity-based screens) represent the first limitation. The second is that, in some extreme environments, cell
densities are so low that the amount of DNA available for cloning is extremely
small, implying additional steps like whole genome amplification. Sequence-based
screens are limited to already known gene families and cannot identify completely
new genes frequent in metagenome databases like CAMERA (Seshadri et al. 2007,
http://camera.calit2.net). Moreover, many genes in metagenome libraries can not be
expressed successfully in organisms such as E. coli and therefore, cannot be detected
by activity-based enzymatic screens. Attempts have been made recently to circumvent these limitations and these have led to the development of innovative methods
like substrate-induced gene expression screening (SIGEX) (Uchiyama et al. 2005).
J. Querellou et al.
screening procedures and directed evolution have been applied with success by several research groups and SMEs as well as larger corporations (Diversa, Genencor,
Degussa, Henkel, etc.) to different targets, enzymes and/or novel natural products.
Table 8.3 gives an overview of the most significant metagenomic contributions
during the recent years.
Though limited to ocean surface microbes, the Sorcerer II/GOS expedition represented a milestone for metagenomics and the team predicted more than six million
proteins in the GOS data, approximately twice the number of proteins present at
that time in the databases (Yooseph et al. 2007). The practical utility of the Sorcerer
II results for marine biotechnology is hard to estimate and their exploitation for
that purpose will take many more years to come to fruition. One of the primary
goals of the Sorcerer II studies was to understand the rate of discoveries of protein
families with the increasing number of protein predictions. From the relationship
observed, they concluded that their sampling of the protein universe is far away
from saturation, meaning that many more protein families remain to be discovered.
This conclusion is subject to controversy and Koonin (Koonin 2007) suggested comparing not only the increase in the number of protein clusters with the number of
sequences observed in the Sorcerer II data but also adding the rate of increase of
orthologs. The virome (viral fraction of the microbiome in a given habitat) of several marine regions has been studied and this has also revealed a completely novel
diversity of proteins (Breitbart et al. 2002, 2004, Angly et al. 2006). The results from
genomics projects dedicated to Archaea viral proteins with unknown functions have
shown that marine viruses represent a reservoir of interest for the discovery of new
folds and novel structures and therefore novel protein families (Vestergaard et al.
2008).
With the advent of metagenomic approaches, the microbial diversity is increasingly accessible and we might expect an exponential growth in the number
of biocatalysts potentially useful for the industry, as already demonstrated by
some recent work on nitrilases, lipases and esterases (Robertson et al. 2004,
Bertram et al. 2008), at least when functional screens are efficient. Note, however, that in contrast to global marine metagenome surveys, these results were
obtained in direct collaboration with industry, with clearly defined biotechnological
goals.
There are two main obstacles to the biotechnological exploitation of marine
extremophiles. Screening methods (sequence-based and activity-based screens) represent the first limitation. The second is that, in some extreme environments, cell
densities are so low that the amount of DNA available for cloning is extremely
small, implying additional steps like whole genome amplification. Sequence-based
screens are limited to already known gene families and cannot identify completely
new genes frequent in metagenome databases like CAMERA (Seshadri et al. 2007,
http://camera.calit2.net). Moreover, many genes in metagenome libraries can not be
expressed successfully in organisms such as E. coli and therefore, cannot be detected
by activity-based enzymatic screens. Attempts have been made recently to circumvent these limitations and these have led to the development of innovative methods
like substrate-induced gene expression screening (SIGEX) (Uchiyama et al. 2005).