Related Experiment Video
Updated: May 25, 2026

A Customizable Approach for the Enzymatic Production and Purification of Diterpenoid Natural Products
Published on: October 4, 2019
Functional expression of a valencene dioxygenase from Pleurotus sapidus in E. coli
Kateryna Zelena1, Ulrich Krings, Ralf G Berger
1Gottfried Wilhelm Leibniz University Hannover, Institute of Food Chemistry, Callinstr. 5, D-30167 Hannover, Germany. kateryna.zelena@lci.uni-hannover.de
Abstract:
Valencene dioxygenase (ValOx) from the edible basidiomycete Pleurotus sapidus converted the sesquiterpene (+)-valencene to the valuable grapefruit flavour (+)-nootkatone and to nootkatols through intermediate hydroperoxides. Expression of the enzyme was carried out in the cytosol and periplasm of Escherichia coli. The heterologous production led to high yields of inclusion bodies. The poor yield of soluble recombinant protein was improved by various strategies including cold shock expression, chaperone co-expression, and employment of mutant E. coli strains. Up to 60 mg of the biologically active, soluble ValOx was produced by cold shock under control of the cspA promoter at 8 °C in the BL21(DE3)Star strain and co-expression of the E. coli trigger factor. The recombinant enzyme, purified using the N-terminal His tag, showed the catalytic properties of the wild-type enzyme, as was confirmed by the LC-MS analysis of hydroperoxide intermediates and GC-MS analysis of the volatile products.
Related Concept Videos
Regioselective Formation of Enolates
Catalytically Perfect Enzymes
Regioselectivity of Electrophilic Additions-Peroxide Effect
Oxidation of Alkenes: Syn Dihydroxylation with Osmium Tetraoxide
Diels–Alder Reaction: Characteristics of Dienophiles
Characteristics of Dienophiles
Generally, the best dienophiles are alkenes containing electron-withdrawing substituents such as carbonyl, nitrile, and nitro groups. The feasibility of a Diels–Alder reaction depends on...
Coordination of Gene Expression Processes in Bacteria

