Diversification of Catalytic Function in a Synthetic Family of Chimeric Cytochrome P450s
Abstract
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.
Additional Information
© 2007 Elsevier Ltd. Under an Elsevier user license. Received 20 December 2006, Accepted 22 January 2007, Available online 23 March 2007. This work is supported by the National Institutes of Health (R01 GM068664-0) and a National Science Foundation Predoctoral Fellowship (to M.C.). The authors thank Sally A. Kim for critically reading the manuscript, Daniela C. Dieterich for help with Figure 2, Figure 4, and Christopher Snow for Figure 5. M.L., C.R.O, and F.H.A. designed and planned the project, and M.L., M.C., C.R.O., and Y.L. performed the experiments. M.C., M.L., and C.R.O. analyzed the data. M.L., M.C., and F.H.A. prepared the manuscript.Attached Files
Accepted Version - nihms20582.pdf
Supplemental Material - 1-s2.0-S1074552107000373-mmc1.pdf
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Additional details
- PMCID
- PMC1991292
- Eprint ID
- 88969
- Resolver ID
- CaltechAUTHORS:20180821-082843212
- NIH
- R01 GM068664-0
- NSF Predoctoral Fellowship
- Created
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2018-08-21Created from EPrint's datestamp field
- Updated
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2021-11-16Created from EPrint's last_modified field