Thermal behaviour and tolerance to ionic liquid [emim]OAc in GH10 xylanase from Thermoascus aurantiacus SL16W

Abstract

GH10 xylanase from Thermoascus aurantiacus strain SL16W (TasXyn10A) showed high stability and activity up to 70-75 °C. The enzyme's half-lives were 101 h, 65 h, 63 min and 6 min at 60, 70, 75 and 80 °C, respectively. The melting point (T m), as measured by DSC, was 78.5 °C, which is in line with a strong activity decrease at 75-80 °C. The biomass-dissolving ionic liquid 1-ethyl-3-methylimidazolium acetate ([emim]OAc) in 30 % concentration had a small effect on the stability of TasXyn10A; T m decreased by only 5 °C. It was also observed that [emim]OAc inhibited much less GH10 xylanase (TasXyn10A) than the studied GH11 xylanases. The K m of TasXyn10A increased 3.5-fold in 15 % [emim]OAc with xylan as the substrate, whereas the approximate level of V max was not altered. The inhibition of enzyme activity by [emim]OAc was lesser at higher substrate concentrations. Therefore, high solid concentrations in industrial conditions may mitigate the inhibition of enzyme activity by ionic liquids. Molecular docking experiments indicated that the [emim] cation has major binding sites near the catalytic residues but in lower amounts in GH10 than in GH11 xylanases. Therefore, [emim] cation likely competes with the substrate when binding to the active site. The docking results indicated why the effect is lower in GH10.

Screening of glycoside hydrolases and ionic liquids for fibre modification

“…In previous studies and in this study, family GH10 xylanases were generally found to be more IL‐tolerant compared with family GH11 enzymes . Earlier findings with [EMIM]AcO suggest that the IL interferes with substrate binding in the active site and this competitive inhibition may be more pronounced with family GH11 xylanases .…”

“…Earlier findings with [EMIM]AcO suggest that the IL interferes with substrate binding in the active site and this competitive inhibition may be more pronounced with family GH11 xylanases . The difference in activity among these enzymes in the presence of ILs appears to be based on differences in the active site and was revealed by molecular docking studies between the enzyme and the IL cation . Probably the narrow and deep active site of GH11 xylanases allows transient binding of large amounts of [EMIM] + cations, thus preventing the binding of a substrate.…”

“…23 In previous studies and in this study, family GH10 xylanases were generally found to be more IL-tolerant compared with family GH11 enzymes. 35 Earlier findings with [EMIM]AcO suggest that the IL interferes with substrate binding in the active site and this competitive inhibition may be more pronounced with family GH11 xylanases. 24 The difference in activity among these enzymes in the presence of ILs appears to be based on differences in the active site and was revealed by molecular docking studies between the enzyme and the IL cation.…”

“…5(A) and (B), respectively). Enzyme tolerance to ILs varied greatly, although the GH10 xylanases (TfXYN10A, TmXYN10B, GH10) showed generally higher IL tolerance 23,35 over the GH11 xylanases (DtXYN11B, GH11). 24 Among the studied enzymes TfXYN10A was the most IL-tolerant enzyme retaining 100% and 48% of relative activity (averaged) in the presence of 15% and 40% IL, respectively.…”

“…24 The difference in activity among these enzymes in the presence of ILs appears to be based on differences in the active site and was revealed by molecular docking studies between the enzyme and the IL cation. 35,36 Probably the narrow and deep active site of GH11 xylanases allows transient binding of large amounts of [EMIM] + cations, thus preventing the binding of a substrate. Consequently, the activity is hindered by competitive inhibition posed by binding of the IL cation.…”