Investigating the applicability of the CYP102A1-decoy-molecule system to other members of the CYP102A subfamily

Cytochrome P450 enzymes (CYPs) have attracted much promise as biocatalysts in a push for cleaner and more environmentally friendly catalytic systems. However, changing the substrate specificity of CYPs, such as CYP102A1, can be a challenging task, requiring laborious mutagenesis. An alternative approach is the use of decoy molecules that “trick” the enzyme into becoming active by impersonating the native substrate. Whilst the decoy molecule system has been extensively developed for CYP102A1, its general applicability for other CYP102-family enzymes has yet to be shown. Herein, we demonstrate that decoy molecules can “trick” CYP102A5 and A7 into becoming active and hydroxylating non-native substrates. Furthermore, significant differences in decoy molecule selectivity as well as decoy molecule binding were observed. The X-ray crystal structure of the CYP102A5 haem domain was solved at 2.8 Å, delivering insight into a potential substate-binding site that differs significantly from CYP102A1. Decoy molecules were shown to be compatible for use with both CYP102A5 and CYP102A7. Both CYP102A5 and CYP102A7 displayed a unique and unexpected decoy molecule preference. The crystal structure of CYP102A5 was solved at a resolution of 2.8 Å. [Display omitted] •Decoy molecules were shown to be compatible for use with both CYP102A5 and A7.•Both CYP102A5 and A7 displayed a unique and unexpected decoy molecule preference.•Decoy molecules were found to bind in a two-step fashion to CYP102A5 and A7 that was not observed for A1.•We report the first structure of CYP102A5 at a resolution of 2.8 Å.•The structure of CYP102A5 reveals a potential binding site that differs significantly from the CYP102A1 reference system.