|Ximenes, Eduardo - Purdue University|
|Kim, Youngmi - Purdue University|
|Mosier, Nathan - Purdue University|
|Ladisch, Michael - Purdue University|
Submitted to: Enzyme and Microbial Technology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/3/2010
Publication Date: 11/1/2010
Citation: Ximenes, E., Kim, Y., Mosier, N., Dien, B.S., Ladisch, M. 2011. Deactivation of cellulases by phenols. Enzyme and Microbial Technology. 48(1):54-60.
Interpretive Summary: Cellulosic ethanol depends upon the treatment of lignocelluloses with enzymes (e.g., cellulases) to produce sugars for fermentation. Prior to saccharifying the plant biomass with enzymes, a thermo-chemical pretreatment is needed to “loosen” up the plant cell wall matrix. During this pretreatment step, additional chemicals are released from the plant cell wall, including a family of chemicals named phenols. In this study, a group of plant cell wall related phenols were examined for their effect on enzymes used for processing biomass. It was found that many of the compounds inactivated cellulases. Tannic acid was especially troublesome in this regard. It was also found that in the case of one of these enzymes, the amount of inhibition could be greatly reduced by substituting an enzyme from a different type of fungus. Results from this paper will aide researchers in engineering enzymes that work better on actual pretreated biomass, eventually reducing enzyme costs associated with converting biomass into biofuels.
Technical Abstract: Pretreatment of lignocellulosic materials may result in the release of inhibitors and deactivators of cellulose enzyme hydrolysis. We report the identification of phenols with major inhibition and/or deactivation effect on enzymes used for conversion of cellulose to ethanol. The inhibition effects were measured by combining the inhibitors (phenols) with enzyme and substrate immediately at the beginning of the assay. The deactivation effects were determined by pre-incubating phenols with cellulases or beta-glucosidases for specified periods of time, prior to the respective enzyme assays. Tannic, gallic, hydroxy-cinnamic, and 4-hydroxybenzoic acids, together with vanillin caused 20–80% deactivation of cellulases and/or beta-glucosidases after 24 h of pre-incubation while enzymes pre-incubated in buffer alone retained all of their activity. The strength of the inhibition or deactivation effect depended on the type of enzyme, the microorganism from which the enzyme was derived, and the type of phenolic compounds present. Beta-glucosidase from Aspergillus niger was the most resistant to inhibition and deactivation, requiring about 5 and 10-fold higher concentrations, respectively, for the same levels of inhibition or deactivation as observed for enzymes from Trichoderma reesei. Of the phenol molecules tested, tannic acid was the single, most damaging aromatic compound that caused both deactivation and reversible loss (inhibition) of all of enzyme activities tested.