Inactivation of Cytochrome P-450 by a Catalytically Generated Cyclobutadiene Species

Article abstract of DOI:10.1021/ja00287a042

2,3-Bis(carbethoxy)-2,3-diazabicyclo<2.2.0>hex-5-ene (DDBCH), synthesized by a route analogues to that reported for the preparation of the bis(carbomethoxy) analogue, inactivates the phenobarbital-inducible cytochrome P-450 isozymes of rat liver in a time-, NADPH-, and oxygen-dependent manner.The enzymes are protected by carbon monoxide and competitive alternative substrates but not by glutathione.The cyclobutenyl ? bond and the diazabicyclo<2.2.0> skeleton are required for destructive activity, but hydrolytic removal of the carbamate groups is not.Enzyme inactivation reflects alkylation of the prosthetic heme group of the enzyme by a catalytically generated reactive species.The alkylated prosthetic group has been isolated and has been characterized, after demetallation and esterification, as N-(2-cyclobutenyl)protoporphyrin IX.The N-alkyl group is primarily located on the nitrogen of pyrrole ring D.DDBCH is thus oxidized by cytochrome P-450 to a cyclobutadienoid species that alkylates the prosthetic heme group.