Previous work has shown that P-450 2B11 is responsible for the unique
ability of dogs to metabolize and eliminate certain highly-chlorinated
biphenyls such as 2,2',4,4',5,5'-hexachlorobiphenyl (245-HCB), wherea
s the related P-450 2B forms in rat and rabbit are unable to metaboliz
e the compound to any significant degree. To determine the structural
basis for this functional diversity, hybrid enzymes were generated. Su
ccess with this approach required a careful choice of second enzyme an
d common substrate with which to assess the functional integrity of th
e hybrid proteins. The choices of P-450 2B5 from rabbit as the second
enzyme and androstenedione as the substrate were based in part on the
finding that P-450 2B11 and P-450 2B5 hydroxylate androstenedione with
similar overall activities but distinct profiles. Enzymatic studies w
ith eight hybrid enzymes provided evidence for two regions of P-450 2B
11 and 2B5, between residues 95-239 and 240-370, that appear to be inv
olved in defining substrate specificity for androstenedione, and three
regions of P-450 2B11, between residues 95-239, 240-370, and 371-494,
that contain amino acids necessary for metabolism of 245-HCB. This de
liberate approach to the creation of hybrid cytochromes P-450 has gene
rated a series of enzymes that will be central to further structure-fu
nction studies of the cytochromes P-450 2B.