RESUMO
An Escherichia coli whole-cell biocatalyst for the direct hydroxylation of benzene to phenol has been developed. By adding amino acid derivatives as decoy molecules to the culture medium, wild-type cytochrome P450BM3 (P450BM3) expressed in E.coli can be activated and non-native substrates hydroxylated, without supplementing with NADPH. The yield of phenol reached 59 % when N-heptyl-l-prolyl-l-phenylalanine (C7-Pro-Phe) was employed as the decoy molecule. It was shown that decoy molecules, especially those lacking fluorination, reached the cytosol of E. coli, thus imparting inâ vivo catalytic activity for the oxyfunctionalisation of non-native substrates to intracellular P450BM3.
Assuntos
Proteínas de Bactérias/metabolismo , Benzeno/metabolismo , Escherichia coli/metabolismo , NADPH-Ferri-Hemoproteína Redutase/metabolismo , Fenóis/metabolismo , Proteínas de Bactérias/genética , Benzeno/química , Biocatálise , Biotransformação , Hidroxilação , NADPH-Ferri-Hemoproteína Redutase/genética , Fenóis/química , Especificidade por SubstratoRESUMO
The selective hydroxylation of benzene to phenol, without the formation of side products resulting from overoxidation, is catalyzed by cytochrome P450BM3 with the assistance of amino acid derivatives as decoy molecules. The catalytic turnover rate and the total turnover number reached 259â min-1 P450BM3-1 and 40 200â P450BM3-1 when N-heptyl-l-proline modified with l-phenylalanine (C7-l-Pro-l-Phe) was used as the decoy molecule. This work shows that amino acid derivatives with a totally different structure from fatty acids can be used as decoy molecules for aromatic hydroxylation by wild-type P450BM3. This method for non-native substrate hydroxylation by wild-type P450BM3 has the potential to expand the utility of P450BM3 for biotransformations.