FAD-dependent enzyme-catalysed intermolecular [4+2] cycloaddition in natural product biosynthesis

Chemoenzymatic Total Syntheses of Artonin I with an Intermolecular Diels–Alderase

“…This indicates the existence of the hydroxy group at C‐4’ position in flavonoid dienes can improve the reactivity of flavonoid dienes toward MaDA, probably via forming hydrogen‐bonding interactions with residues N357 and N374 in MaDA. [ 22 ]…”

“…Gene coding truncated N terminus MaDA without signal peptide was cloned into pI-secSUMOstar vector, expressed using the Bac-to-Bac system (Invitrogen, USA), and purified using the Ni-NTA resin (GE healthcare, USA) according to the previously reported protocol. [22] The purified protein contains a SUMO tag in the N terminal. Standard curve of artonin I (1) was determined for quantification, the data were shown in Figure S1, Supporting Information.…”

“…Similar with the proposed biosynthetic pathway of other Diels–Alder type natural products in Morus plants, [ 21 ] the enzyme‐catalyzed oxidation (or dehydrogenation) and Diels–Alder reaction were proposed as the final two key steps in the biosynthesis of artonin I by exogenous feeding of diene precursor in the Morus alba cell cultures. [ 18 ] Recently, we have identified the first standalone and intermolecular Diels–Alderase (MaDA) from the M. alba cell cultures, which catalyzes the endo‐ and enantio‐specific Diels–Alder reaction to generate chalcomoracin and other related Diels–Alder type natural products, [ 22 ] demonstrating the synthetic utility of MaDA in the chemo‐enzymatic synthesis of endo‐selective Diels–Alder type natural products. Thus, we envision that the MaDA‐catalyzed Diels–Alder reaction may provide an effective means to produce other Diels–Alder type natural products such as artonin I with high endo‐ and enantioselectivity, which has been proven to be challenging in chemical catalysis.…”

Biosynthetic Intermediate Probes for Visualizing and Identifying the Biosynthetic Enzymes of Plant Metabolites

“…Affinity‐based probes are widely used in chemical proteomics approaches for target identification of bioactive molecules [19] . Recently, we successfully identified an intermolecular Diels‐Alderase [20] in Morus alba by using a BIP with a photoaffinity group, thus demonstrating that BIP‐based target identification is an alternative way to identify missing enzymes in plant natural product biosynthesis (Figure 1B). Moreover, when combining transcriptome analysis and BIP‐based target identification, candidate genes in both methods will be highly reliable for biochemical characterization.…”

Mechanistic Study of Hetero-Diels–Alder [4 + 2] Cycloaddition Reactions Between 2-Nitro-1H-Pyrrole and Isoprene

“…The cycloadditions [4 + 2] of Diels-Alder (DA reaction) is important for the synthesis of natural products and biologically active molecules [1][2][3][4][5][6][7]. Research on this reaction in order to develop new synthesis methods and simultaneously improve yields, regioselectivity and stereoselectivity is constantly developing [8,9].…”