Distinct Mechanisms of Acetic Acid Inhibition in Thermophilic GH10 and GH11 Xylanases

Acetic acid generated during dilute acid or hydrothermal pretreatment of lignocellulosic biomass often acts as a potent inhibitor of glycoside hydrolases (GHs), including xylanases, thereby reducing the efficiency of xylan saccharification. This lowers xylose and xylooligosaccharide yields and overall carbon recovery in lignocellulosic biorefineries. In this study, we compared the inhibition patterns of a GH10 xylanase (TcrGH10) and a GH11 xylanase (TsaGH11) by acetic acid. Both enzymes exhibited only a modest loss of activity (~10-15%) between 0 and 0.8% acetic acid but were almost completely inactivated at 1.0%, suggesting a critical concentration range. Global fitting of kinetic data at 0, 0.3, and 0.75% acetic acid to inhibition models indicated mixed-type inhibition with a predominant non-competitive component for TcrGH10, whereas TsaGH11 predominantly followed an uncompetitive inhibition pattern. These findings elucidate family-specific inhibition behaviors and establish a critical acetic acid threshold, offering guidance for process optimization and the development of acetic acid-tolerant xylanases.