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44 3 Monooxygenase-Catalyzed Redox Cascade Biotransformations

As depicted in Scheme 3.1, a reducing agent (Donor H ) activates molecular
2
oxygen, which is then transferred onto the substrate. When employing a monooxy-
genase, the second oxygen is reduced to water; this process requires two electrons,
which are delivered by a cofactor (NADH or NADPH). Biocatalytic oxygenation
of various compounds is a reaction performed mainly by heme, non-heme iron,
copper pterin, and flavin-dependent enzymes [1]. Monooxygenases are typically
highly chemo-, regio-, and/or enantioselective, making them highly attractive in
biocatalysis [2, 3]. In this chapter, the focus will be on monooxygenases exclusively,
providing an overview on their applicability in multistep catalysis.

Mono-oxygenases
Sub + Donor H 2 + O 2 Cofactor-recycling SubO + Donor + H 2 O

Scheme 3.1 Illustration of molecular oxygen fixation by monooxygenases.
Several reviews have been published recently describing the general concept
and the advantages of cascade or domino reactions [4]. Their potential power lies
in the capability to overcome thermodynamic hurdles in multistep syntheses [5].
Different designs of cascade processes have been described in the literature [6,
7]. One approach is represented by a linear cassette of processes, such as the
well-known glycolytic metabolic pathway in nature. Orthogonal cascades are a
complementary concept, and they are related to the transformation of a substrate
into the product thanks to the regeneration of cofactors or co-substrates via
coupled reactions. A classic example is represented by nicotine amide-dependent
monooxygenases that are coupled to a cofactor-regenerating enzyme (formate
dehydrogenase or glucose dehydrogenase). The third strategy is closely related to
the orthogonal principle and is depicted as parallel cascades where two substrates
are converted into two products by two distinct biocatalytic entities. In contrast to
the orthogonal approach, one in which both products are valuable has a higher
economic value. The last type is the cyclic cascade, where one out of a mixture of
substrates is converted into an intermediate, which is then transformed back to
the starting materials. Dynamic kinetic resolutions of racemic starting materials
represent one example of the latter class. All concepts are quite appealing especially
in the field of redox biocatalysis [8–11]. Enzyme cascades can be carried out in
the manifolds of biological systems by exploiting or engineering their metabolic
networks for catalysis, or, alternatively, completely new pathways can be assembled
that are independent of the host’s metabolism. Additionally, in vitro cascades can be
performed by using either isolated enzymes or cell-free extracts. Both approaches
have their advantages and disadvantages, problems and challenges, which will be
illustrated in the following sections [12].

3.1.3
Effective Cofactor Recycling

From the very beginning, scientists active in the field of biocatalysis wanted
to promote the use of enzymes as powerful homogeneous catalysts for organic