Spiro-Ring Formation is Catalyzed by a Multifunctional Dioxygenase in Austinol Biosynthesis

Abstract

Austinol, a fungal meroterpenoid derived from 3,5-dimethylorsellinic acid, has a unique chemical structure with a remarkable spiro-lactone ring system. Despite the recent identification of its biosynthetic gene cluster and targeted gene-deletion experiments, the process for the conversion of protoaustinoid A (2), the first tetracyclic biosynthetic intermediate, to the spiro-lactone preaustinoid A3 (7) has remained enigmatic. Here we report the mechanistic details of the enzyme-catalyzed, stereospecific spiro-lactone ring-forming reaction, which is catalyzed by a non-heme iron-dependent dioxygenase, AusE, along with two flavin monooxygenases, the 5'-hydroxylase AusB and the Baeyer-Villiger monooxygenase AusC. Remarkably, AusE is a multifunctional dioxygenase that is responsible for the iterative oxidation steps, including the oxidative spiro-ring-forming reaction, to produce the austinol scaffold.

3,5‐Dimethylorsellinic Acid Derived Meroterpenoids from Eupenicillium sp. 6A‐9, a Fungus Isolated from the Marine Sponge Plakortis simplex

“…[30] Notably, the completely elucidated pathways for austinol [21,22,27] and berkeleydione, [29] for which the known compounds preaustinoid A1 (7) and preaustinoid A3 (10) acquired here are biosynthetic intermediates, enabled us to propose detailed mechanisms for the biosynthesis of the five related new meroterpenoids 1-5 (Scheme 1).…”

“…The biosynthesis of intriguing meroterpenoids metabolized by Aspergillus and Penicillium has been investigated for decades, and recently the molecular bases (biosynthetic genes/enzymes) for five 3,5-dimethylorsellinic acid (DMOA) and farnesyl pyrophosphate derived meroterpenoids, andrastin A, austinol, terretonin, anditomin, and berkeleydione, have been reported [20][21][22][23][24][25][26][27][28][29] and thoroughly reviewed. [30] Notably, the completely elucidated pathways for austinol [21,22,27] and berkeleydione, [29] for which the known compounds preaustinoid A1 (7) and preaustinoid A3 (10) acquired here are biosynthetic intermediates, enabled us to propose detailed mechanisms for the biosynthesis of the five related new meroterpenoids 1-5 (Scheme 1).…”

“…Next, 7 is subjected to two successive oxidations in which 7 undergoes dehydrogenation to preaustinoid A2 (14) and oxidative rearrangement to spiro-lactone 10, which is known to be catalyzed by a multifunctional Fe II /α-KG-dependent dioxygenase, AusE. [27] Apparently, two steps involving ester hydrolyzation and methyl esterification of 7 give rise to 1-methoxy-hydropreaustinoid A1 (3). Meanwhile, as reported previously, [29] 7 is transformed into berkeleydione (16) via berkeleyone B (15) in two consecutive oxidations, also catalyzed by a multifunctional Fe II /α-KG-dependent dioxygenase, PrhA.…”

Oxidative Cyclization in Natural Product Biosynthesis

“…574 The epoxide-mediated formation of the meroterpenoid scaffold was discussed previously in Scheme 100 (Section 4.3). Starting with 951 , generation of the bicyclo[2.2.2]octane ring system in andiconin ( 956 ) is catalyzed by AndA (Scheme 136).…”

Enantioselective α-Hydroxylation of β-Keto Esters Catalyzed by Cinchona Alkaloid Derivatives

“…8 Moreover, (S)-methyl 5-chloro-2-hydroxy-1-oxo-2,3-dihydro-1H-indene-2-carboxylate is an important intermediate in the synthesis of the insecticide Indoxacarb (Scheme 1) which was developed by the DuPont company. 9 Direct α-hydroxylation of β-keto esters is the most convenient approach to obtain the α-hydroxy β-dicarbonyl units.…”