Purification and characterization of the sesquiterpene cyclase trichodiene synthetase from Fusarium sporotrichioides.
The sesquiterpene cyclase, trichodiene synthetase, has been purified from a supernatant fraction of Fusarium sporotrichioides by hydrophobic interaction, anion exchange, and gel filtration chromatography. Purified enzyme had a specific activity 15-fold higher than that previously reported for preparations of terpene cyclases. Molecular weight determinations by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration chromatography indicated the enzyme to be a dimer with a subunit of Mr 45,000. The requirement of Mg2+ (Km 0.1 mM) for activity could be partially substituted with Mn2+ at a concentration of 0.01 mM, but higher concentrations of Mn2+ were inhibitory. Maximum activity was observed between pH 6.75 and pH 7.75. The Km for farnesyl pyrophosphate was 0.065 microM.
D019755:Carbon-Carbon Lyases; D002845:Chromatography / Q000379:methods; D005670:Fusarium / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D008190:Lyases / Q000302:isolation & purification* / Q000378:metabolism; D008670:Metals / Q000494:pharmacology; D013379:Substrate Specificity
D008670:Metals; D008190:Lyases; D019755:Carbon-Carbon Lyases; C051130:trichodiene synthetase