| Abstract
| - Norcarane, bicyclo[4.1.0]heptane, has been widely used as a mechanistic probe in studies of oxidationscatalyzed by several iron-containing enzymes. We report here that, in addition to oxygenated products,norcarane is also oxidized by iron-containing enzymes in desaturase reactions that give 2-norcarene and3-norcarene. Furthermore, secondary products from further oxidation reactions of the norcarenes areproduced in yields that are comparable to those of the minor products from oxidation of the norcarane.We studied oxidations catalyzed by a representative spectrum of iron-containing enzymes including fourcytochrome P450 enzymes, CYP2B1, CYPΔ2B4, CYPΔ2E1, and CYPΔ2E1 T303A, and three diironenzymes, soluble methane monooxygenase (sMMO) from Methylococcus capsulatus (Bath), toluenemonooxygenase (ToMO) from Pseudomonas stutzeri OX1, and phenol hydroxylase (PH) fromPseudomonas stutzeri OX1. 2-Norcarene and 3-norcarene and their oxidation products were found in allreaction mixtures, accounting for up to half of the oxidation products in some cases. In total, more than20 oxidation products were identified from the enzyme-catalyzed reactions of norcarane. The putativeradical-derived product from the oxidation of norcarane, 3-hydroxymethylcyclohexene (21), and theputative cation-derived product from the oxidation of norcarane, cyclohept-3-enol (22), coelute with otheroxidation products on low-polarity GC columns. The yields of product 21 found in this study are smallerthan those previously reported for the same or similar enzymes in studies where the products from norcareneoxidations were ignored, and therefore, the limiting values for lifetimes of radical intermediates producedin the enzyme-catalyzed oxidation reactions are shorter than those previously reported.
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