Structural insights of RmXyn10A – A prebiotic-producing GH10 xylanase with a non-conserved aglycone binding regionстатья
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Аннотация:Hydrolysis of arabinoxylan (AX) by glycoside hydrolase family 10 (GH10) xylanases produces xylo- and arabinoxylo-
oligosaccharides ((A)XOS) which have shown prebiotic effects. The thermostable GH10 xylanase
RmXyn10A has shown great potential to produce (A)XOS. In this study, the structure of RmXyn10A was investigated,
the catalytic module by homology modelling and site-directed mutagenesis and the arrangement of
its five domains by small-angle X-ray scattering (SAXS). Substrate specificity was explored in silico by manual
docking and molecular dynamic simulations. It has been shown in the literature that the glycone subsites of
GH10 xylanases are well conserved and our results suggest that RmXyn10A is no exception. The aglycone
subsites are less investigated, and the modelled structure of RmXyn10A suggests that loop β6α6 in the aglycone
part of the active site contains a non-conserved α-helix, which blocks the otherwise conserved space of subsite
+2. This structural feature has only been observed for one other GH10 xylanase. In RmXyn10A, docking revealed
two alternative binding regions, one on either side of the α-helix. However, only one was able to accommodate
arabinose-substitutions and the mutation study suggests that the same region is responsible for
binding XOS. Several non-conserved structural features are most likely to be responsible for providing affinity for
arabinose-substitutions in subsites +1 and +2. The SAXS rigid model of the modular arrangement of RmXyn10A
displays the catalytic module close to the cell-anchoring domain while the carbohydrate binding modules are
further away, likely explaining the observed lack of contribution of the CBMs to activity.