86
H. Zhu et al.
the production of cytochalasin H (7) in M. grisea P450 knockout strains (ΔpyiD and
ΔpyiG) in a combinatorial biosynthesis study [55].
The above information supports the evolution analysis of ACE cluster in M. oryzae
conducted by Wolfe and Bradshaw’s groups, which suggested four genes, the PKSNRPS, the trans-ER, the pDA and the α,β-hydrolase gene, as the core set of genes
(Fig. 21) involved in the production of a group of secondary metabolites that are
modified differentially by other accessory enzymes [250, 261]. Using these core
genes, several ACE1 clusters homologated cryptic gene clusters were discovered
in other filamentous fungi (Fig. 21g), although the metabolites they encode remain
unknown [2, 250, 261].
4.2 Investigations into the Biosynthesis of Cytochalasans
Before the characterization of the che gene cluster in P. expansum, there was hardly
any information available on the molecular basis of cytochalasan biosynthesis. Even
at the present time, the function of hybrid PKS-NRPS in fungi [254–257], which
play a core role in cytochalasan clusters, has not been known very long. Using
RNA-mediated gene silencing experiments, Hertweck and coworkers characterized
the functions of two core genes (cheA and cheB) in the che cluster and proposed
a cytochalasan biosynthetic pathway, in that the PKS part of the hybrid synthase
CheA and the enoyl reductase CheB act together to synthesize a polyketide chain.
Subsequently, this is passed on to the NRPS and condensed with an activated tryptophan [253], and then catalyzed by a C-terminal reductase domain in CheA, with
a reductive off-loading then taking place to release an aminoaldehyde intermediate
that can further form a pyrrolinone via a Knoevenagel condensation. This pyrrolinone can function as a dienophile, and could undergo intramolecular Diels–Alder
(DA) cyclization with the polyketide terminal diene to generate the isoindolone-fused
macrocycle (Scheme 1). The release mechanism and the formation of the isoindolonefused macrocycle was also proposed in an alternative way that a functional domain
within the PKS–NRPS acted as a Dieckmann cyclase, as observed in (aspyr-)idone
and tenellin biosynthesis [256, 257], and directly releases a tetramic acid derivative
[253], which further undergoes a series of subsequent reactions such as dehydration,
intramolecular DA cyclization and reduction. The roles of the oxidases CheD, CheE
and CheG in this cluster were proposed as modifying the cytochalasan backbones to
yield chaetoglobosins A (210) and C (212) (Scheme 1).
To date, several cytochalasan gene clusters have been identified, and the individual
steps to form the cytochalasan backbones and structural diversity were partially
characterized. As described in a previous review by Skellam [2], the biosynthesis
of cytochalasans may be divided into three stages: the early stages, the formation of
the linear polyketide conjunct pyrrolinone; the middle stages, the formation of the
macrocycle fused isoindolone via DA cyclization, and the late stages that lead to
modifications on the cytochalasan framework.
H. Zhu et al.
the production of cytochalasin H (7) in M. grisea P450 knockout strains (ΔpyiD and
ΔpyiG) in a combinatorial biosynthesis study [55].
The above information supports the evolution analysis of ACE cluster in M. oryzae
conducted by Wolfe and Bradshaw’s groups, which suggested four genes, the PKSNRPS, the trans-ER, the pDA and the α,β-hydrolase gene, as the core set of genes
(Fig. 21) involved in the production of a group of secondary metabolites that are
modified differentially by other accessory enzymes [250, 261]. Using these core
genes, several ACE1 clusters homologated cryptic gene clusters were discovered
in other filamentous fungi (Fig. 21g), although the metabolites they encode remain
unknown [2, 250, 261].
4.2 Investigations into the Biosynthesis of Cytochalasans
Before the characterization of the che gene cluster in P. expansum, there was hardly
any information available on the molecular basis of cytochalasan biosynthesis. Even
at the present time, the function of hybrid PKS-NRPS in fungi [254–257], which
play a core role in cytochalasan clusters, has not been known very long. Using
RNA-mediated gene silencing experiments, Hertweck and coworkers characterized
the functions of two core genes (cheA and cheB) in the che cluster and proposed
a cytochalasan biosynthetic pathway, in that the PKS part of the hybrid synthase
CheA and the enoyl reductase CheB act together to synthesize a polyketide chain.
Subsequently, this is passed on to the NRPS and condensed with an activated tryptophan [253], and then catalyzed by a C-terminal reductase domain in CheA, with
a reductive off-loading then taking place to release an aminoaldehyde intermediate
that can further form a pyrrolinone via a Knoevenagel condensation. This pyrrolinone can function as a dienophile, and could undergo intramolecular Diels–Alder
(DA) cyclization with the polyketide terminal diene to generate the isoindolone-fused
macrocycle (Scheme 1). The release mechanism and the formation of the isoindolonefused macrocycle was also proposed in an alternative way that a functional domain
within the PKS–NRPS acted as a Dieckmann cyclase, as observed in (aspyr-)idone
and tenellin biosynthesis [256, 257], and directly releases a tetramic acid derivative
[253], which further undergoes a series of subsequent reactions such as dehydration,
intramolecular DA cyclization and reduction. The roles of the oxidases CheD, CheE
and CheG in this cluster were proposed as modifying the cytochalasan backbones to
yield chaetoglobosins A (210) and C (212) (Scheme 1).
To date, several cytochalasan gene clusters have been identified, and the individual
steps to form the cytochalasan backbones and structural diversity were partially
characterized. As described in a previous review by Skellam [2], the biosynthesis
of cytochalasans may be divided into three stages: the early stages, the formation of
the linear polyketide conjunct pyrrolinone; the middle stages, the formation of the
macrocycle fused isoindolone via DA cyclization, and the late stages that lead to
modifications on the cytochalasan framework.