PDBsum entry: 1w2u

Hydrolase

PDB id: 1w2u

Contents

Protein chain

224 a.a. *

Ligands

BGC-BGC-BGC-BGC

SO4

PG4

Waters ×257

* Residue conservation analysis

PDB id:

1w2u

Name:

Hydrolase

Title:

X-ray crystal structure of the catalytic domain of humicola grisea cel12a in complex with a soaked thio cellotetraose

Structure:

Endoglucanase. Chain: a. Fragment: catalytic domain residues 31-254. Engineered: yes. Other_details: the crystal structure is a complex with a soaked cellopentaose. The fifth glucose unit is, however, not visible in the electron density

Source:

Humicola grisea. Organism_taxid: 5527. Expressed in: aspergillus niger. Expression_system_taxid: 5061

Resolution:

1.52Å R-factor: 0.144 R-free: 0.179

Authors:

G.I.Berglund,A.Shaw,J.Stahlberg,L.Kenne,T.H.Driguez, C.Mitchinson,M.Sandgren

Key ref:

M.Sandgren et al. (2004). Crystal complex structures reveal how substrate is bound in the -4 to the +2 binding sites of Humicola grisea Cel12A. J Mol Biol, 342, 1505-1517. PubMed id: 15364577 DOI: 10.1016/j.jmb.2004.07.098

Date:

08-Jul-04 Release date: 16-Sep-04

Protein chain

Q8NJY3  (Q8NJY3_9ASCO) -  Endoglucanase

Seq: 254 a.a.

Struc: 224 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

 Gene Ontology (GO) functional annotation 

• Biological process: polysaccharide catabolic process (1 term)
• Biochemical function: hydrolase activity, hydrolyzing O-glycosyl compounds (2 terms)

DOI no: 10.1016/j.jmb.2004.07.098

J Mol Biol 342:1505-1517 (2004)

PubMed id: 15364577

Crystal complex structures reveal how substrate is bound in the -4 to the +2 binding sites of Humicola grisea Cel12A.

M.Sandgren, G.I.Berglund, A.Shaw, J.Ståhlberg, L.Kenne, T.Desmet, C.Mitchinson.

ABSTRACT

As part of an ongoing enzyme discovery program to investigate the properties and catalytic mechanism of glycoside hydrolase family 12 (GH 12) endoglucanases, a GH family that contains several cellulases that are of interest in industrial applications, we have solved four new crystal structures of wild-type Humicola grisea Cel12A in complexes formed by soaking with cellobiose, cellotetraose, cellopentaose, and a thio-linked cellotetraose derivative (G2SG2). These complex structures allow mapping of the non-covalent interactions between the enzyme and the glucosyl chain bound in subsites -4 to +2 of the enzyme, and shed light on the mechanism and function of GH 12 cellulases. The unhydrolysed cellopentaose and the G2SG2 cello-oligomers span the active site of the catalytically active H.grisea Cel12A enzyme, with the pyranoside bound in subsite -1 displaying a S31 skew boat conformation. After soaking in cellotetraose, the cello-oligomer that is found bound in site -4 to -1 contains a beta-1,3-linkage between the two cellobiose units in the oligomer, which is believed to have been formed by a transglycosylation reaction that has occurred during the ligand soak of the protein crystals. The close fit of this ligand and the binding sites occupied suggest a novel mixed beta-glucanase activity for this enzyme.

Selected figure(s)

Figure 2.
Figure 2. Electron density maps for the H. grisea Cel12A-tetrasaccharide (b), cellopentaose (c) and cellobiose (d) complexes. The electron density for the combined Cel12A-cellohexaose complex (a) is a theoretical combination of the structures from the tetrasaccharide complex ( -4 to -2 glucans), and that from the cellopentaose complex ( -1 to +2 glucans). The maps shown are maximum-likelihood s[A]-weighted 2|F[obs]| -|F[calc]| maps, contoured at 1s. The electron density has been cut around the cellooligomers using a masking radius of 1.5 Å, for clarity. The catalytic nucleophile (Glu120) and the acid/base (Glu205) are shown for reference.

Figure 5.
Figure 5. Overlay of the covalently bound 2-deoxy-2-fluoro-cellotrioside ligand (coloured in red), bound in sites -3, -2 and -1 of S. lividans Cel12A,12 with the mixed b-1,3-1,4-tetraose ligand (coloured in gold) bound in sites -4 to -2 of H. grisea Cel12A. The catalytic nucleophile (Glu120) and the acid/base (Glu205) of H. grisea Cel12A are drawn for reference. The overlay shows how the mixed b-1,3-1,4-tetraose ligand extends along the bottom of the cleft in H. grisea Cel12A, whereas a regular b-1,4-glucan chain would extend upwards from the bottom of the cleft.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2004, 342, 1505-1517) copyright 2004.

Figures were selected by an automated process.