DNA shuffling of cytochrome P450 enzymes

Abstract

DNA family shuffling is an ef ficient method for creating libraries of novel enzymes, in which a high proportion of mutants exhibit correct folding and possess catalytic properties distinct from the starting material. The evolutionary arrangement of cytochromes P450 into subfamilies of enzymes with highly similar nucleotide sequences but distinct catalytic properties renders them excellent starting material for DNA family shuf fling experiments. This chapter provides a general method for creating libraries of shuf fled P450s from two or more related sequences and incorporates several recent improvements to previously published methods. © Springer Science+Business Media New York 2013.