PDBsum entry 1y9m

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protein ligands links
Hydrolase PDB id
Protein chain
517 a.a. *
NAG ×3
GOL ×4
Waters ×639
* Residue conservation analysis
PDB id:
Name: Hydrolase
Title: Crystal structure of exo-inulinase from aspergillus awamori spacegroup p212121
Structure: Exo-inulinase. Chain: a. Fragment: sequence database residues 20-537. Ec:
Source: Aspergillus awamori. Organism_taxid: 105351. Strain: 2250
1.89Å     R-factor:   0.167     R-free:   0.200
Authors: R.A.P.Nagem,A.L.Rojas,A.M.Golubev,O.S.Korneeva,E.V.Eneyskaya A.A.Kulminskaya,K.N.Neustroev,I.Polikarpov
Key ref:
R.A.Nagem et al. (2004). Crystal structure of exo-inulinase from Aspergillus awamori: the enzyme fold and structural determinants of substrate recognition. J Mol Biol, 344, 471-480. PubMed id: 15522299 DOI: 10.1016/j.jmb.2004.09.024
16-Dec-04     Release date:   28-Dec-04    
Go to PROCHECK summary

Protein chain
Pfam   ArchSchema ?
Q96TU3  (Q96TU3_ASPAW) -  Extracellular exo-inulinase inuE
537 a.a.
517 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: E.C.  - Fructan beta-fructosidase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]

      Reaction: Hydrolysis of terminal, non-reducing 2,1- and 2,6-linked beta-D-fructofuranose residues in fructans.
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     extracellular region   1 term 
  Biological process     metabolic process   3 terms 
  Biochemical function     hydrolase activity     4 terms  


DOI no: 10.1016/j.jmb.2004.09.024 J Mol Biol 344:471-480 (2004)
PubMed id: 15522299  
Crystal structure of exo-inulinase from Aspergillus awamori: the enzyme fold and structural determinants of substrate recognition.
R.A.Nagem, A.L.Rojas, A.M.Golubev, O.S.Korneeva, E.V.Eneyskaya, A.A.Kulminskaya, K.N.Neustroev, I.Polikarpov.
Exo-inulinases hydrolyze terminal, non-reducing 2,1-linked and 2,6-linked beta-d-fructofuranose residues in inulin, levan and sucrose releasing beta-d-fructose. We present the X-ray structure at 1.55A resolution of exo-inulinase from Aspergillus awamori, a member of glycoside hydrolase family 32, solved by single isomorphous replacement with the anomalous scattering method using the heavy-atom sites derived from a quick cryo-soaking technique. The tertiary structure of this enzyme folds into two domains: the N-terminal catalytic domain of an unusual five-bladed beta-propeller fold and the C-terminal domain folded into a beta-sandwich-like structure. Its structural architecture is very similar to that of another member of glycoside hydrolase family 32, invertase (beta-fructosidase) from Thermotoga maritima, determined recently by X-ray crystallography The exo-inulinase is a glycoprotein containing five N-linked oligosaccharides. Two crystal forms obtained under similar crystallization conditions differ by the degree of protein glycosylation. The X-ray structure of the enzyme:fructose complex, at a resolution of 1.87A, reveals two catalytically important residues: Asp41 and Glu241, a nucleophile and a catalytic acid/base, respectively. The distance between the side-chains of these residues is consistent with a double displacement mechanism of reaction. Asp189, which is part of the Arg-Asp-Pro motif, provides hydrogen bonds important for substrate recognition.
  Selected figure(s)  
Figure 4.
Figure 4. (a) Stereoview of fructose:enzyme interactions in the catalytic site of exo-inulinase. The fructose and the neighboring residues are shown. An omit electron density map (mF[obs] -DF[calc], calc), contoured at 3s, is displayed around the catalytic site of EI. (b) Schematic design of the interactions between the active-site residues and the fructose molecule (in pink). Hydrogen bonds are shown as cyan dotted lines. The Figure was drawn using the program LIGPLOT.56
Figure 5.
Figure 5. A representation of a double displacement mechanism of reaction with an overall retention of the configuration at the anomeric carbon atom of the substrate, catalysed by exo-inulinase from Aspergillus awamori. The R group may represent fructose units of inulin.
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2004, 344, 471-480) copyright 2004.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
  21418142 A.Bujacz, M.Jedrzejczak-Krzepkowska, S.Bielecki, I.Redzynia, and G.Bujacz (2011).
Crystal structures of the apo form of β-fructofuranosidase from Bifidobacterium longum and its complex with fructose.
  FEBS J, 278, 1728-1744.
PDB codes: 3pig 3pij
20563857 M.A.Rodríguez, O.F.Sánchez, and C.J.Alméciga-Díaz (2011).
Gene cloning and enzyme structure modeling of the Aspergillus oryzae N74 fructosyltransferase.
  Mol Biol Rep, 38, 1151-1161.  
21082263 S.Li, Y.Yan, Z.Zhou, H.Yu, Y.Zhan, W.Zhang, M.Chen, W.Lu, S.Ping, and M.Lin (2011).
Single amino acid residue changes in subsite -1 of levansucrase from Zymomonas mobilis 10232 strongly influence the enzyme activities and products.
  Mol Biol Rep, 38, 2437-2443.  
19936386 A.Homann, and J.Seibel (2009).
Chemo-enzymatic synthesis and functional analysis of natural and modified glycostructures.
  Nat Prod Rep, 26, 1555-1571.  
19622545 C.S.Kim, B.Pierre, M.Ostermeier, L.L.Looger, and J.R.Kim (2009).
Enzyme stabilization by domain insertion into a thermophilic protein.
  Protein Eng Des Sel, 22, 615-623.  
19088319 D.Linde, I.Macias, L.Fernández-Arrojo, F.J.Plou, A.Jiménez, and M.Fernández-Lobato (2009).
Molecular and biochemical characterization of a beta-fructofuranosidase from Xanthophyllomyces dendrorhous.
  Appl Environ Microbiol, 75, 1065-1073.  
19514896 K.Vijayaraghavan, D.Yamini, V.Ambika, and N.S.Sowdamini (2009).
Trends in inulinase production--a review.
  Crit Rev Biotechnol, 29, 67-77.  
19726634 L.Schroeven, W.Lammens, A.Kawakami, M.Yoshida, A.Van Laere, and W.Van den Ende (2009).
Creating S-type characteristics in the F-type enzyme fructan:fructan 1-fructosyltransferase of Triticum aestivum L.
  J Exp Bot, 60, 3687-3696.  
19127444 T.Mutanda, B.Wilhelmi, and C.G.Whiteley (2009).
Controlled production of fructose by an exoinulinase from Aspergillus ficuum.
  Appl Biochem Biotechnol, 159, 65-77.  
18821076 T.Zhang, F.Gong, Z.Chi, G.Liu, Z.Chi, J.Sheng, J.Li, and X.Wang (2009).
Cloning and characterization of the inulinase gene from a marine yeast Pichia guilliermondii and its expression in Pichia pastoris.
  Antonie Van Leeuwenhoek, 95, 13-22.  
19129163 W.Lammens, K.Le Roy, L.Schroeven, A.Van Laere, A.Rabijns, and W.Van den Ende (2009).
Structural insights into glycoside hydrolase family 32 and 68 enzymes: functional implications.
  J Exp Bot, 60, 727-740.  
19122997 Z.Chi, Z.Chi, T.Zhang, G.Liu, and L.Yue (2009).
Inulinase-expressing microorganisms and applications of inulinases.
  Appl Microbiol Biotechnol, 82, 211-220.  
18366639 G.Meng, and K.Fütterer (2008).
Donor substrate recognition in the raffinose-bound E342A mutant of fructosyltransferase Bacillus subtilis levansucrase.
  BMC Struct Biol, 8, 16.
PDB codes: 3byj 3byk 3byl 3byn
17963237 J.Mátrai, W.Lammens, A.Jonckheer, K.Le Roy, A.Rabijns, W.Van den Ende, and M.De Maeyer (2008).
An alternate sucrose binding mode in the E203Q Arabidopsis invertase mutant: an X-ray crystallography and docking study.
  Proteins, 71, 552-564.
PDB code: 2oxb
17293485 C.Goosen, X.L.Yuan, J.M.van Munster, A.F.Ram, M.J.van der Maarel, and L.Dijkhuizen (2007).
Molecular and biochemical characterization of a novel intracellular invertase from Aspergillus niger with transfructosylating activity.
  Eukaryot Cell, 6, 674-681.  
17849258 E.Ricca, V.Calabrò, S.Curcio, and G.Iorio (2007).
The state of the art in the production of fructose from inulin enzymatic hydrolysis.
  Crit Rev Biotechnol, 27, 129-145.  
17888113 K.Le Roy, M.Verhaest, A.Rabijns, S.Clerens, A.Van Laere, and W.Van den Ende (2007).
N-glycosylation affects substrate specificity of chicory fructan 1-exohydrolase: evidence for the presence of an inulin binding cleft.
  New Phytol, 176, 317-324.  
17335500 M.Verhaest, W.Lammens, K.Le Roy, C.J.De Ranter, A.Van Laere, A.Rabijns, and W.Van den Ende (2007).
Insights into the fine architecture of the active site of chicory fructan 1-exohydrolase: 1-kestose as substrate vs sucrose as inhibitor.
  New Phytol, 174, 90.
PDB codes: 2add 2ade 2aey 2aez
17502268 S.Moriyama, and K.Ohta (2007).
Functional characterization and evolutionary implication of the internal 157-amino-acid sequence of an exoinulinase from Penicillium sp. strain TN-88.
  J Biosci Bioeng, 103, 293-297.  
17192265 W.S.Jung, C.K.Hong, S.Lee, C.S.Kim, S.J.Kim, S.I.Kim, and S.Rhee (2007).
Structural and functional insights into intramolecular fructosyl transfer by inulin fructotransferase.
  J Biol Chem, 282, 8414-8423.
PDB codes: 2inu 2inv
16899050 L.K.Ozimek, S.Kralj, T.Kaper, M.J.van der Maarel, and L.Dijkhuizen (2006).
Single amino acid residue changes in subsite -1 of inulosucrase from Lactobacillus reuteri 121 strongly influence the size of products synthesized.
  FEBS J, 273, 4104-4113.  
17139091 M.Verhaest, W.Lammens, K.Le Roy, B.De Coninck, C.J.De Ranter, A.Van Laere, W.Van den Ende, and A.Rabijns (2006).
X-ray diffraction structure of a cell-wall invertase from Arabidopsis thaliana.
  Acta Crystallogr D Biol Crystallogr, 62, 1555-1563.
PDB code: 2ac1
17018033 T.Ritsema, L.Hernández, A.Verhaar, D.Altenbach, T.Boller, A.Wiemken, and S.Smeekens (2006).
Developing fructan-synthesizing capability in a plant invertase via mutations in the sucrose-binding box.
  Plant J, 48, 228-237.  
  16511152 M.Verhaest, K.Le Roy, S.Sansen, B.De Coninck, W.Lammens, C.J.De Ranter, A.Van Laere, W.Van den Ende, and A.Rabijns (2005).
Crystallization and preliminary X-ray diffraction study of a cell-wall invertase from Arabidopsis thaliana.
  Acta Crystallogr Sect F Struct Biol Cryst Commun, 61, 766-768.  
16041066 R.A.Nagem, A.L.Ambrosio, A.L.Rojas, M.V.Navarro, A.M.Golubev, R.C.Garratt, and I.Polikarpov (2005).
Getting the most out of X-ray home sources.
  Acta Crystallogr D Biol Crystallogr, 61, 1022-1030.  
16158237 T.Ritsema, A.Verhaar, I.Vijn, and S.Smeekens (2005).
Using natural variation to investigate the function of individual amino acids in the sucrose-binding box of fructan:fructan 6G-fructosyltransferase (6G-FFT) in product formation.
  Plant Mol Biol, 58, 597-607.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB codes are shown on the right.