Selective aliphatic carbon–hydrogen bond activation of protected alcohol substrates by cytochrome P450 enzymes

Abstract

Protected cyclohexanol and cyclohex-2-enol substrates, containing benzyl ether and benzoate ester moieties, were designed to fit into the active site of the Tyr96Ala mutant of cytochrome P450cam. The protected cyclohexanol substrates were efficiently and selectively hydroxylated by the mutant enzyme at the trans C-H bond of C-4 on the cyclohexyl ring. The selectivity of oxidation of the benzoate ester protected cyclohexanol could be altered by making alternative amino acid substitutions in the P450cam active site. The addition of the double bond in the cyclohexyl ring of the benzoate ester protected cyclohex-2-enol has a debilitative effect on the activity of the Tyr96Ala mutant with this substrate. However, the Phe87Ala/Tyr96Phe double mutant, which introduces space at a different location in the active site than the Tyr96Ala mutant, was able to efficiently hydroxylate the C-H bonds of 1-cyclohex-2-enyl benzoate at the allylic C-4 position. Mutations at Phe87 improved the selectivity of the oxidation of 1-phenyl-1-cyclohexylethylene to trans-4-phenyl-ethenylcyclohexanol (92%) when compared to single mutants at Tyr96 of P450cam.

Midazolam as a Probe for Heterotropic Drug-Drug Interactions Mediated by CYP3A4

“…The position of the substrate and SOM preferences may be changed by mutations [ 67 , 68 , 69 ] and by interactions with allosteric effectors [ 6 , 26 , 44 , 70 ]. Vice versa, concentration-dependent changes in the SOM ratio due to the presence of various compounds can be used as a direct indication of ability of these compounds to serve as allosteric effectors.…”

Biocatalytic Oxidation Reactions: A Chemist's Perspective

“…[26] Similarly, Bell and co-workers achieved the selective 4-hydroxylation of ester-o re ther-protected cyclohexanols. [27] Very impressive changes in the regioselectivity of aP 450 monooxygenasecatalysed hydroxylation of macrolides have been achieved with this approach (Scheme 8). [28] Aside from the selectivity issues,P 450 monooxygenase reactions also suffer from the generally low turnover numbers of the oxyfunctionalisation catalysts.…”

“…Early contributions by Griengl and co‐workers demonstrated that the hydroxylation of cyclopentanone does not occur using the P450 monooxygenase from Beauveria bassiana whereas the corresponding spirooxazolidine was converted rather efficiently . Similarly, Bell and co‐workers achieved the selective 4‐hydroxylation of ester‐ or ether‐protected cyclohexanols . Very impressive changes in the regioselectivity of a P450 monooxygenase‐catalysed hydroxylation of macrolides have been achieved with this approach (Scheme )…”

Biokatalytische Oxidationsreaktionen – aus der Sicht eines Chemikers

“…In ihren bahnbrechenden Arbeiten konnten Griengl und Mitarbeiter zeigen, dass die entsprechenden Spirooxazolidine allerdings sehr gut umgesetzt werden . Neuere Arbeiten aus dem Arbeitskreis Bell erweitern dieses Konzept auf die selektive 4‐Hydroxylierung Ether‐ oder Ester‐geschützter Cyclohexanole …”