Approaches to random mutagenesis with cytochrome P450 and glutathione transferase enzymes

A general method was developed for the selection of cytochrome P450 mutants from randomized libraries on the basis of catalytic function. The approach utilizes the ability of P450 expressed in Escherichia coli (with its reduc tase) to catalyze the N-hydroxylation of heterocyclic amines, yielding muta gens that can be detected using a lacZ screen. This system can be used, in principle, to screen mutants of any enzyme that activates chemicals to muta gens and can be expressed in bacteria. The mammalian theta class glutathion e (GSH) transferases (GST) (rat 5-5, human T1-1) and methylophilus DM11 act ivate dihaloethanes and dihalomethanes to mutagens and may be amenable to t his approach. (C) 2001 Elsevier Science Ireland Ltd. All rights reserved.