PDBsum entry 1h49

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Hydrolase PDB id
Protein chains
491 a.a. *
Waters ×491
* Residue conservation analysis
PDB id:
Name: Hydrolase
Title: Crystal structure of the inactive double mutant of the maize beta-glucosidase zmglu1-e191d-f198v in complex with dimboa-glucoside
Structure: Beta-glucosidase. Chain: a, b. Fragment: beta-glucosidase, residues 55-566. Synonym: chloroplast precursor, gentiobiase, cellobiase, beta-d-glucoside glucohydrolase, glu1. Engineered: yes. Mutation: yes. Other_details: in complex with the ligand dimboa-glucoside
Source: Zea mays. Maize. Organism_taxid: 4577. Strain: cv. Mutin. Tissue: coleoptile. Cell: plys s cells. Expressed in: escherichia coli. Expression_system_taxid: 562.
1.90Å     R-factor:   0.185     R-free:   0.221
Authors: M.Czjzek,J.Moriniere,L.Verdoucq,D.R.Bevan,B.Henrissat, A.Ese
Key ref:
L.Verdoucq et al. (2003). Mutational and structural analysis of aglycone specificity in maize and sorghum beta-glucosidases. J Biol Chem, 278, 25055-25062. PubMed id: 12684498 DOI: 10.1074/jbc.M301978200
25-Feb-03     Release date:   11-Mar-03    
Go to PROCHECK summary

Protein chains
Pfam   ArchSchema ?
P49235  (BGLC_MAIZE) -  4-hydroxy-7-methoxy-3-oxo-3,4-dihydro-2H-1,4-benzoxazin-2-yl glucoside beta-D-glucosidase 1, chloroplastic
566 a.a.
491 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class 2: E.C.  - 4-hydroxy-7-methoxy-3-oxo-3,4-dihydro-2H-1,4-benzoxazin-2-yl glucoside
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
1. (2R)-4-hydroxy-7-methoxy-3-oxo-3,4-dihydro-2H-1,4-benzoxazin-2-yl beta-D-glucopyranoside + H2O = 2,4-dihydroxy-7-methoxy-2H-1,4- benzoxazin-3(4H)-one + D-glucose
2. (2R)-4-hydroxy-3-oxo-3,4-dihydro-2H-1,4-benzoxazin-2-yl beta-D- glucopyranoside + H2O = 2,4-dihydroxy-2H-1,4-benzoxazin-3(4H)-one + D-glucose
(2R)-4-hydroxy-7-methoxy-3-oxo-3,4-dihydro-2H-1,4-benzoxazin-2-yl beta-D-glucopyranoside
+ H(2)O
= 2,4-dihydroxy-7-methoxy-2H-1,4- benzoxazin-3(4H)-one
+ D-glucose
(2R)-4-hydroxy-3-oxo-3,4-dihydro-2H-1,4-benzoxazin-2-yl beta-D- glucopyranoside
+ H(2)O
= 2,4-dihydroxy-2H-1,4-benzoxazin-3(4H)-one
+ D-glucose
   Enzyme class 3: E.C.  - Beta-glucosidase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Hydrolysis of terminal, non-reducing beta-D-glucose residues with release of beta-D-glucose.
Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     plastid   2 terms 
  Biological process     metabolic process   3 terms 
  Biochemical function     hydrolase activity     4 terms  


DOI no: 10.1074/jbc.M301978200 J Biol Chem 278:25055-25062 (2003)
PubMed id: 12684498  
Mutational and structural analysis of aglycone specificity in maize and sorghum beta-glucosidases.
L.Verdoucq, M.Czjzek, J.Moriniere, D.R.Bevan, A.Esen.
Plant beta-glucosidases display varying substrate specificities. The maize beta-glucosidase isozyme Glu1 (ZmGlu1) hydrolyzes a broad spectrum of substrates in addition to its natural substrate DIMBOA-Glc (2-O-beta-d-glucopyranosyl-4-hydroxy-7-methoxy-1,4-benzoxaxin-3-one), whereas the sorghum beta-glucosidase isozyme Dhr1 (SbDhr1) hydrolyzes exclusively its natural substrate dhurrin (p-hydroxy-(S)-mandelonitrile-beta-d-glucoside). Structural data from cocrystals of enzyme-substrate and enzyme-aglycone complexes have shown that five amino acid residues (Phe198, Phe205, Trp378, Phe466, and Ala467) are located in the aglycone-binding site of ZmGlu1 and form the basis of aglycone recognition and binding, hence substrate specificity. To study the mechanism of substrate specificity further, mutant beta-glucosidases were generated by replacing Phe198, Phe205, Asp261, Met263, Phe377, Phe466, Ala467, and Phe473 of Glu1 by Dhr1 counterparts. The effects of mutations on enzyme activity and substrate specificity were studied using both natural and artificial substrates. The simple mutant replacing Phe198 by a valine had the most drastic effect on activity, because the capacity of this enzyme to hydrolyze beta-glucosides was almost completely abolished. The analysis of this mutation was completed by a structural study of the double mutant ZmGlu1-E191D,F198V in complex with the natural substrate. The structure reveals that the single mutation F198V causes a cascade of conformational changes, which are unpredictable by standard molecular modeling techniques. Some other mutations led to drastic effects: replacing Asp261 by an asparagine decreases the catalytic efficiency of this simple mutant by 75% although replacing Tyr473 by a phenylalanine increase its efficiency by 300% and also provides a new substrate specificity by hydrolyzing dhurrin.
  Selected figure(s)  
Figure 2.
FIG. 2. Diagrammatic representation of wild-type parental ZmGlu1 and SbDhr1 isozymes and 2 chimeras generated by domain swapping, as well as 8 site-directed mutants. Sequences of the swapped regions are given on the top of the diagram in Chim16 and Chim21. The sites where ZmGlu1 and SbDhr1 differ within the swapped regions are highlighted in red, as are the amino acids exchanged by site-directed mutagenesis.
Figure 3.
FIG. 3. Close-up view of the aglycone recognition and binding site of ZmGlu1-E191D,F198V in complex with DIMBOA-glucoside. a, final electron density of ZmGlu1-E191D,F198V. The map is a SIGMAA weighted 2F[obs] - F[calc] synthesis contoured at 1 , showing the well defined density around the residues undergoing a positional shift in this mutant structure. b, the initial difference Fourier map (F[obs] - F[calc]) located in the active site pocket, contoured at a 2 level, showing the presence of a substrate molecule. c, structural superimposition of ZmGlu1-E191D (in blue) and ZmGlu1-E191D, F198V (the residues are color coded with standard atom-type colors), both in complex with DIMBOA-Glc (omitted from this figure). The F198V mutation (highlighted in blue) induces a positional change of Phe^205, Phe^466 takes a different rotameric conformation, and Glu464 in the glycone binding pocket is also displaced. d, the same structural superimposition as in c, viewed in a different orientation, illustrating the drastic change in the position of the DIMBOA-glucoside molecule in ZmGlu1-E191D,F198V (red substrate molecule and atom type colors for the residues) with respect to ZmGlu1-E191D (dark blue, substrate molecule, and blue, residues).
  The above figures are reprinted by permission from the ASBMB: J Biol Chem (2003, 278, 25055-25062) copyright 2003.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20490603 J.R.Ketudat Cairns, and A.Esen (2010).
  Cell Mol Life Sci, 67, 3389-3405.  
19056785 F.S.Kittur, H.Y.Yu, D.R.Bevan, and A.Esen (2009).
Homolog of the maize beta-glucosidase aggregating factor from sorghum is a jacalin-related GalNAc-specific lectin but lacks protein aggregating activity.
  Glycobiology, 19, 277-287.  
18615662 A.D.Hill, and P.J.Reilly (2008).
Computational analysis of glycoside hydrolase family 1 specificities.
  Biopolymers, 89, 1021-1031.  
18422657 L.M.Mendonça, and S.R.Marana (2008).
The role in the substrate specificity and catalysis of residues forming the substrate aglycone-binding site of a beta-glycosidase.
  FEBS J, 275, 2536-2547.  
19016858 R.Dopitová, P.Mazura, L.Janda, R.Chaloupková, P.Jerábek, J.Damborský, T.Filipi, N.S.Kiran, and B.Brzobohatý (2008).
Functional analysis of the aglycone-binding site of the maize beta-glucosidase Zm-p60.1.
  FEBS J, 275, 6123-6135.  
18023045 T.Tsukada, K.Igarashi, S.Fushinobu, and M.Samejima (2008).
Role of subsite +1 residues in pH dependence and catalytic activity of the glycoside hydrolase family 1 beta-glucosidase BGL1A from the basidiomycete Phanerochaete chrysosporium.
  Biotechnol Bioeng, 99, 1295-1302.  
18033585 J.Stöckigt, and S.Panjikar (2007).
Structural biology in plant natural product biosynthesis--architecture of enzymes from monoterpenoid indole and tropane alkaloid biosynthesis.
  Nat Prod Rep, 24, 1382-1400.  
16233798 E.Ishikawa, T.Sakai, H.Ikemura, K.Matsumoto, and H.Abe (2005).
Identification, cloning, and characterization of a Sporobolomyces singularis beta-galactosidase-like enzyme involved in galacto-oligosaccharide production.
  J Biosci Bioeng, 99, 331-339.  
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.