DNA-Mediated Assembly of Cytochrome P450 BM3 Subdomains
| Autor: | Christof M. Niemeyer, Michael Erkelenz, Chi-Hsien Kuo |
|---|---|
| Rok vydání: | 2011 |
| Předmět: |
Recombinant Fusion Proteins
Allosteric regulation Protein Engineering 010402 general chemistry 01 natural sciences Biochemistry Catalysis chemistry.chemical_compound Colloid and Surface Chemistry Allosteric Regulation Bacterial Proteins Cytochrome P-450 Enzyme System DNA nanotechnology NADPH-Ferrihemoprotein Reductase chemistry.chemical_classification Bioconjugation Bacteria 010405 organic chemistry Oligonucleotide DNA General Chemistry Protein engineering Protein Structure Tertiary 0104 chemical sciences Enzyme chemistry Drug delivery Biophysics Laurates |
| Zdroj: | Journal of the American Chemical Society |
| ISSN: | 1520-5126 0002-7863 |
| DOI: | 10.1021/ja204993s |
| Popis: | Cytochrome P450 BM3 is a versatile enzyme, which holds great promise for applications in biocatalysis and biomedicine. We here report on the generation of a hybrid DNA-protein device based on the two subdomains of BM3, the reductase domain BMR and the porphyrin domain BMP. Both subdomains were fused genetically to the HaloTag protein, a self-labeling enzyme, allowing for the bioconjugation with chloroalkane-modified oligonucleotides. The subdomain-DNA-chimeras could be reassembled by complementary oligonucleotides, thus leading to reconstitution of the monooxygenase activity of BM3 holoenzyme, as demonstrated by conversion of the reporter substrate 12-pNCA. Arrangement of the two chimeras on a switchable DNA scaffold allowed one to control the distance between both subdomains, as indicated by the DNA-dependent activity of the holoenzyme. Furthermore, a switchable chimeric device was constructed, in which monooxygenase activity could be turned off by DNA strand displacement. This study demonstrates that P450 BM3 engineering and strategies of DNA nanotechnology can be merged to open up novel ways for the development of novel screening systems or responsive catalysts with potential applications in drug delivery. |
| Databáze: | OpenAIRE |
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