Location: Bioproducts ResearchTitle: Divalent metal activation of a GH43 ß-xylosidase) Author
Submitted to: Enzyme and Microbial Technology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/26/2012
Publication Date: 2/5/2013
Citation: Lee, C.C., Braker, J.D., Grigorescu, A.A., Wagschal, K.C., Jordan, D.B. 2013. Divalent metal activation of a GH43 ß-xylosidase. Enzyme and Microbial Technology. 52(2):84-90. Interpretive Summary: Hemicellulose (xylan) is the second largest component of lignocellulosic biomass. As such, it is critical that this substrate be fully utilized to ensure the economic viability of biorefineries. Xylan is a polymer that must be hydrolyzed into the simple sugar xylose which can then be fermented by microorganisms into fuel and other chemical feedstocks. The ß-xylosidase enzymes cleave more chemical bonds in this process than any other enzyme and thus play an essential role in the breakdown of xylan. We now report the discovery of a new, highly-active ß-xylosidase enzyme. We describe the effects of divalent metal cations upon the enzyme activity. This is the first detailed report of metal effects upon this category of enzyme, and the results have important implications of how industry should optimize the reaction conditions for these enzymes.
Technical Abstract: Depolymerization of xylan, a major fraction of lignocellulosic biomass, releases fermentable xylose which can be converted into transportation fuels and chemical feedstocks. One of the requisite enzymes for the breakdown of xylan is ß-xylosidase. A gene encoding the 324-amino acid ß-xylosidase, RS223-BX, was cloned from an anaerobic mixed microbial culture. This glycoside hydrolase belongs to family 43. It can be activated by Ca2+, Co2+, Mg2+, Mn2+ and Ni2+ (e.g., 27-fold by Mg2+). Sedimentation equilibrium centrifugation experiments indicated that the divalent metal cations mediate multimerization of the enzyme from a dimeric to a tetrameric state, which have equal catalytic activity on an active-site basis. Compared to the determined active sites of other GH43 ß-xylosidases, the predicted active site of RS223-BX contains two additional amino acids with carboxylated sidechains that provide a place for divalent metal cations to reside. Thus, the divalent metal cations likely occupy the active site and participate in the catalytic mechanism. RS223-BX accepts as substrate xylobiose, arabinobiose, 4-nitrophenyl-ß-D-xylopyranoside, and 4-nitrophenyl-a-L-arabinofuranoside. Additionally, the enzyme has good pH and temperature stabilities and a large Ki for D-glucose (1.3 M), favorable properties for performance in saccharification reactors.