The substrate specificity in the O-demethylation of 4-alkylguaiacols by cytochrome P450 AgcAP450

Sónia F. G. Santos, Rajesh Bommareddy, Gary Black, Warispreet Singh*, Meilan Huang*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

1 Citation (Scopus)


Alkylguaiacols are lignin-derived products obtained by reductive catalytic fractionation (RCF) of lignocellulosic biomass. The recently discovered AgcAP450 enzyme from the CYP255A1 family oxidizes a range of 4-alkylguaiacols, having a preference for bulkier substrates such as 4-propylguaiacol over small substrates such as guaiacol. The substrate specificity of AgcAP450 for 4-propylguaiacol, 4-ethylguaiacol, 4-methylguaiacol, and guaiacol was studied using molecular dynamics (MD) simulations. Our results showed that AgcAP450 adopts an open conformation of the substrate access channel, allowing a water flux into the active site, and therefore resulting in a bigger pocket to accommodate bulkier substrates. MD simulations disclosed that the smaller substrates could not stably bind in the catalytic site, and thus it is unlikely for them to be transformed. Further, our quantum mechanics molecular mechanics (QM/MM) study shows that the activation energy barrier for the rate limiting step of the O-demethylation, i.e. hydrogen atom abstraction, is the lowest for AgcAP450 in complex with 4-propylguaicol, compared to that of the enzyme in complex with the other smaller substrates. This unravels the structural feature of the lignin degradation enzyme responsible for the preference toward bulky substrates, and hence sets a basis for engineering enzymes to be exploited for the sustainable conversion of lignocellulosic biomass.
Original languageEnglish
Pages (from-to)2070-2079
Number of pages10
JournalCatalysis Science and Technology
Issue number7
Early online date28 Feb 2023
Publication statusPublished - 7 Apr 2023

Cite this