Background: Degradation of the plant cell wall requires the synergistic action of a consortium of predominantly modular enzymes. In Clostridiae, these biocatalysts are organized into a supramolecular assembly termed a "cellulosome." This multienzyme complex possesses, in addition to its well-described cellulolytic activity, an apparatus specific for xylan degradation. Cinnamic acid esterases hydrolyze the ferulate groups involved in the crosslinking of arabinoxylans to lignin and thus play a key role in the degradation of the plant cell wall in addition to having promising industrial and medical applications. Results: We have cloned and overexpressed the feruloyl esterase module from a 5 domain xylanase, Xyn10B from Clostridium thermocellum. The native structure at 1.6 Å resolution has been solved with selenomethionine multiple wavelength anomalous dispersion and refined to a final Rfree of 17.8%. The structure of a hydrolytically inactive mutant, S954A, in complex with the reaction product ferulic acid has been refined at a resolution of 1.4 Å with an Rfree of 16.0%. Conclusions: The C. thermocellum Xyn10B ferulic acid esterase displays the α/β-hydrolase fold and possesses a classical Ser-His-Asp catalytic triad. Ferulate esterases are characterized by their specificity, and the active center reveals the binding site for ferulic acid and related compounds. Ferulate binds in a small surface depression that possesses specificity determinants for both the methoxy and hydroxyl ring substituents of the substrate. There appears to be a lack of specificity for the xylan backbone, which may reflect the intrinsic chemical heterogeneity of the natural substrate.
Prates, J. A. M., Tarbouriech, N., Charnock, S. J., Fontes, C. M. G. A., Ferreira, L. M. A., & Davies, G. J. (2001). The structure of the feruloyl esterase module of xylanase 10B from Clostridium thermocellum provides insights into substrate recognition. Structure, 9(12), 1183–1190. https://doi.org/10.1016/S0969-2126(01)00684-0