Diversification of catalytic function in a synthetic family of chimeric cytochrome p450s.
We report initial characterization of a synthetic family of more than 3000 cytochrome P450s made by SCHEMA recombination of 3 bacterial CYP102s. A total of 16 heme domains and their holoenzyme fusions with each of the 3 parental reductase domains were tested for activity on 11 different substrates. The results show that the chimeric enzymes have acquired significant functional diversity, including the ability to accept substrates not accepted by the parent enzymes. K-means clustering analysis of the activity data allowed the enzymes to be classified into five distinct groups based on substrate specificity. The substrates can also be grouped such that one can be a "surrogate" for others in the group. Fusion of a functional chimeric heme domain with a parental reductase domain always reconstituted a functional holoenzyme, indicating that key interdomain interactions are conserved upon reductase swapping.
Landwehr
Marco
M
Division of Chemistry and Chemical Engineering, California Institute of Technology, Mail Code 210-41, Pasadena, CA 91125, USA.
Carbone
Martina
M
Otey
Christopher R
CR
Li
Yougen
Y
Arnold
Frances H
FH
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002384:Catalysis; D003001:Cloning, Molecular; D016000:Cluster Analysis; D017124:Conserved Sequence; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D014644:Genetic Variation; D007700:Kinetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D003577:Cytochrome P-450 Enzyme System