Toluene monooxygenase-catalyzed epoxidation of alkenes

Several toluene monooxygenase-producing organisms were tested for their abi lity to oxidize linear alkenes and chloroalkenes three to eight carbons lon g. Each of the wild-type organisms degraded all of the alkenes that were te sted. Epoxides were produced during the oxidation of butene, butadiene, and pentene but not hexene or octadiene. A strain of Escherichia coli expressi ng the cloned toluene-4-monooxygenase (T4MO) of Pseudomanas mendocina KR1 w as able to oxidize butene, butadiene, pentene, and hexene but not octadiene , producing epoxides from all of the substrates that were oxidized. A T4MO- deficient variant of P. mendocina KR1 oxidized alkenes that were five to ei ght carbons long, but no epoxides were detected, suggesting the presence of multiple alkene-degrading enzymes in this organism. The alkene oxidation r ates varied widely (ranging from 0.01 to 033 mu mol of substrate/min/mg of cell protein) and were specific for each organism-substrate pair. The enant iomeric purity of the epoxide products also varied widely, ranging from 54 to >90% of a single epoxide enantiomer. In the absence of more preferred su bstrates, such as toluene or alkenes, the epoxides underwent further toluen e monooxygenase-catalyzed transformations, forming products that were not i dentified.