Page 380 - Cascade_Biocatalysis_Integrating_Stereoselective_and_Environmentally_Friendly_Reactions

356 16 Aldolases as Catalyst for the Synthesis of Carbohydrates and Analogs

aldolases. They are also valuable synthetic tools for the preparation of naturally
occurring compounds and their analogs.
Many attempts have been made during the last two decades to demonstrate
the utility and limitations of aldolases, as illustrated by the plethora of reports
surveyed in the literature. Protein engineering techniques and screening of novel
aldol activities in nature are important tools for future aldolase development,
which is expected to produce unprecedented innovative structures and access to
new reactions. These advances offer promising perspectives for the application
of aldolases, particularly after the thrilling emergence of organocatalysis as an
alternative to biocatalysis.
Aldolase redesign by protein engineering is mainly focused on enhancing the
acceptor and donor tolerance toward a wide structural spectrum of nucleophiles
and electrophiles. In this connection, FSA-like enzymes constitute an attractive
platform for modification because of their intrinsic tolerance toward structurally
diverse donors.
The alteration of the stereochemistry of the newly formed C–C bonds, which is
usually dependent on the substrate structure, is a more difficult task. Precedents
to direct, independent modification of the stereochemistry of the two stereogenic
centers for one single substrate in aldolases have not been reported yet. Stereocom-
plementary aldolases may be reached by directed evolution and rational redesign
[37]. The knowledge of the three-dimensional structure of aldolases will facilitate
further redesigns. In addition, analysis of the distinct tolerances and stereochem-
ical outcome of the native as well as the genetically modified aldolases is helping
in gaining insight into the substrate recognition and orientation in the active site.
The discovery of the precise mechanism of the enzymatic aldol addition process in
combination with the elucidation of enzyme structure is also of paramount impor-
tance. All this knowledge, along with rational and random protein engineering,
is essential for future aldolase development particularly directed to optimize their
activity when combined with several other biocatalysts into a multistep biocatalytic
cascade reactions.

Acknowledgments

The work developed in our research group was supported by the Spanish MICINN
CTQ2012-31605, Generalitat de Catalunya (2009 SGR 00281), and ESF project
COST CM0701.

References

1. (a) Mahrwald, R. (2004) Modern Aldol Modern Aldol Reactions: Metal Cataly-
Reactions: Enolates, Organocatalysis, sis, vol. 2, Wiley-VCH Verlag GmbH,
Biocatalysis and Natural Product Synthe- Weinheim. (c) Evans, D.A., Nelson,
sis,vol.1,Wiley-VCHVerlagGmbH, J.V., and Taber, T.R. (1982) Top. Stere-
Weinheim. (b) Mahrwald, R. (2004) ochem., 13, 1–115. (d) Mukherjee, S.,