GENETICALLY-ENGINEERED BACTERIAL-CELLS AND APPLICATIONS

Heterologous expression systems have proven to be very important in P4 50 research, permitting investigation of many P450s identified only by recombinant (cloning) technologies. They also make it possible to stu dy human P450s since tissue sources for naturally occurring enzymes ar e difficult to obtain. A variety of different heterologous systems hav e been applied to the study of P450s and each one has its own unique a dvantages. For the study of biophysical properties and structure/funct ion relationships, E. coli has proven to be a particularly good system . Both microsomal and mitochondrial P450s can be expressed to high lev els in E. coli and subsequently purified to homogeneity for detailed a nalysis. Techniques of both site directed mutagenesis and random chime ragenesis are very facile in bacteria providing excellent opportunity to analyze specific aspects of structure/function relationships of P45 0s. Furthermore microsomal P450s are active in intact E. coli, activit ies being supported by flavodoxin and flavodoxin reductase, providing the opportunity to develop bioreactors expressing designer P450s. The salient features of P450 expression in bacteria are summarized herein.