Harnessing enzyme plasticity for the synthesis of oxygenated sesquiterpenoids

Abstract

8-Methoxy-γ-humulene, (E)-8-methoxy-β-farnesene, 12-methoxy-β-sesquiphellandrene and 12-methoxyzingiberene can be synthesised in amorphadiene synthase-catalysed reactions from 8- and 12-methoxyfarnesyl diphosphates due to the highly plastic yet tightly controlled carbocationic chemistry of this sesquiterpene cyclase.

Rerouting and Improving Dauc‐8‐en‐11‐ol Synthase from Streptomyces venezuelae to a High Yielding Biocatalyst

“…We and others have recently shown that additional methyl groups can also enzymatically be incorporated into terpenes from methylated IPP derivatives, but in these cases the additional methyl groups were not placed in positions that lead to an altered reactivity of the obtained methylated oligoprenyl diphosphate derivatives. [40,41] Also functional groups such as the keto group in the FPP analogue 16 are tolerated by DcS, similar to recent findings for other terpene synthases, [42][43][44][45] yielding products 33 and 34 with functionalised side chains that allow further chemical transformations. We have also investigated the effect of mutating aromatic amino acid residues that are, according to a homology model, located on the surface of DcS.…”

Modular Chemoenzymatic Synthesis of Terpenes and their Analogues

“…Analogues of terpene synthase substrates have been pivotal to determining the mechanisms of terpene cyclizations by establishing the sequence of carbocation migrations, hydride and methyl shifts and the position of any proton eliminations . These analogues can often act as substrates, forming modified terpene products . Point mutations of terpene synthases can dramatically alter product distributions, catalytic efficiencies, and substrate preferences allowing access to a plethora of new, non‐natural terpenoid products .…”

“…[19][20][21][22] These analogues can often act as substrates, forming modified terpene products. [22][23][24][25] Point mutations of terpene synthases can dramatically alter product distributions, catalytic efficiencies, and substrate preferences allowing access to a plethora of new, non-natural terpenoid products. [26][27][28][29] For example, in addition to farnesyl diphosphate (FDP), (S)-germacrene D synthase (ScGDS) accepts 14,15-dimethyl-FDP as a substrate and cyclizes it to (S)-14,15-dimethylgermacrene D. Strikingly, while the natural product (S)-germacrene D is a repellent of aphids, (S)-14,15-dimethylgermacrene D acts as an attractant.…”

Synthesis, Enzymatic Peptide Incorporation, and Applications of Diazirine-Containing Isoprenoid Diphosphate Analogues

“…Since 7b contains two alkenes with comparable reactivity, careful chromatographic separation of the two isomeric products was necessary to isolate 8b in high purity; these isomeric products have been previously characterized. 12 Overoxidation of 8b to the corresponding aldehyde is also a problem in this type of reaction; hence, a reduction step using NaBH 4 was added following oxidation to increase the yield of the desired alcohol product. Next, 8b was esterified with 5 using DIC to provide the masked product 9b, which was deprotected to yield compound 10b.…”

“…To evaluate the diphosphate probes as substrates for FTase and GGTase-I, a continuous spectrofluorimetric assay was employed that monitors the time-dependent increase in fluorescence that occurs when a dansylated peptide is prenylated. The well-characterized peptide substrates Dns-GCVLS (12) and Dns-GCVLL (13) were used to evaluate the rat FTase or yeast FTase (rFTase or yFTase) 13 and rat GGTase-I (rGGTase) 14 activity, respectively. Incubation of rFTase or yFTase with 1 or 2 revealed efficient use of 1 to yield the prenylated product 14a (Figure S1A and S1B and Figure 2).…”