Isotopic labeling of the heme cofactor in cytochrome p450 and other heme proteins

Abstract

A recombinant bacterial expression system that generates ^sup 13^C-labeled heme or ^sup 15^N-labeled heme in functional cytochrome P450 enzymes and other heme-containing systems is reported here using a mutant strain of Escherichia coli (HU227) in which the HemA gene is inactive. By synthesizing several isotopomers of aminolevulinic acid with ^sup 13^C or ^sup 15^N at different locations, isotopes have been incorporated with high abundance into the heme cofactor of five different cytochrome P450 isoforms, along with one peroxidase. Confirmed both ^sup 13^C- and ^sup 15^N-incorporation; spectral and catalytic assays show the labeled enzymes produced in this system are functional.[PUBLICATION ABSTRACT]