First crystal structure of an endo-inulinase, INU2, from Aspergillus ficuum: Discovery of an extra-pocket in the catalytic domain responsible for its endo-activity

J. Pouyez, A. Mayard, A.-M. Vandamme, G. Roussel, E.A. Perpète, J. Wouters, I. Housen, C. Michaux

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Endo-inulinase is a member of glycosidase hydrolase family 32 (GH32) degrading fructans of the inulin type with an endo-cleavage mode and is an important class of industrial enzyme. In the present study, we report the first crystal structure of an endo-inulinase, INU2, from Aspergillus ficuum at 1.5 Å. It was solved by molecular replacement with the structure of exo-inulinase as search model. The 3D structure presents a bimodular arrangement common to other GH32 enzymes: a N-terminal 5-fold β-propeller catalytic domain with four β-sheets and a C-terminal β-sandwich domain organized in two β-sheets with five β-strands. The structural analysis and comparison with other GH32 enzymes reveal the presence of an extra pocket in the INU2 catalytic site, formed by two loops and the conserved motif W-M(I)-N-D(E)-P-N-G. This cavity would explain the endo-activity of the enzyme, the critical role of Trp40 and particularly the cleavage at the third unit of the inulin(-like) substrates. Crystal structure at 2.1 Å of INU2 complexed with fructosyl molecules, experimental digestion data and molecular modelling studies support these hypotheses.
Original languageEnglish
Pages (from-to)2423-2430
Number of pages8
Issue number11
Publication statusPublished - 1 Nov 2012



  • Endo-inulase
  • Glycoside hydrolase
  • crystal structure
  • Enzyme mechanism
  • Molecular modelling

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