PDBsum entry 1x0c

Hydrolase

PDB id: 1x0c

Contents

Protein chains

549 a.a. *

Ligands

NAG ×23

Waters ×1273

* Residue conservation analysis

PDB id:

1x0c

Name:

Hydrolase

Title:

Improved crystal structure of isopullulanase from aspergillus niger atcc 9642

Structure:

Isopullulanase. Chain: a, b. Engineered: yes

Source:

Aspergillus niger. Organism_taxid: 5061. Expressed in: pichia pastoris. Expression_system_taxid: 4922.

Resolution:

1.70Å R-factor: 0.195 R-free: 0.229

Authors:

M.Mizuno,T.Tonozuka,A.Yamamura,Y.Miyasaka,H.Akeboshi,S.Kamitori, A.Nishikawa,Y.Sakano

Key ref:

M.Mizuno et al. (2008). Crystal structure of Aspergillus niger isopullulanase, a member of glycoside hydrolase family 49. J Mol Biol, 376, 210-220. PubMed id: 18155243 DOI: 10.1016/j.jmb.2007.11.098

Date:

17-Mar-05 Release date: 13-Jun-06

Protein chains

O00105  (IPUA_ASPNG) -  Isopullulanase from Aspergillus niger

Seq: 564 a.a.

Struc: 549 a.a.*

* PDB and UniProt seqs differ at 4 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.2.1.57 - isopullulanase.
• Reaction: Hydrolysis of pullulan to isopanose (6-alpha-maltosylglucose).

DOI no: 10.1016/j.jmb.2007.11.098

J Mol Biol 376:210-220 (2008)

PubMed id: 18155243

Crystal structure of Aspergillus niger isopullulanase, a member of glycoside hydrolase family 49.

M.Mizuno, A.Koide, A.Yamamura, H.Akeboshi, H.Yoshida, S.Kamitori, Y.Sakano, A.Nishikawa, T.Tonozuka.

ABSTRACT

An isopullulanase (IPU) from Aspergillus niger ATCC9642 hydrolyzes alpha-1,4-glucosidic linkages of pullulan to produce isopanose. Although IPU does not hydrolyze dextran, it is classified into glycoside hydrolase family 49 (GH49), major members of which are dextran-hydrolyzing enzymes. IPU is highly glycosylated, making it difficult to obtain its crystal. We used endoglycosidase H(f) to cleave the N-linked oligosaccharides of IPU, and we here determined the unliganded and isopanose-complexed forms of IPU, both solved at 1.7-A resolution. IPU is composed of domains N and C joined by a short linker, with electron density maps for 11 or 12 N-acetylglucosamine residues per molecule. Domain N consists of 13 beta-strands and forms a beta-sandwich. Domain C, where the active site is located, forms a right-handed beta-helix, and the lengths of the pitches of each coil of the beta-helix are similar to those of GH49 dextranase and GH28 polygalacturonase. The entire structure of IPU resembles that of a GH49 enzyme, Penicillium minioluteum dextranase (Dex49A), despite a difference in substrate specificity. Compared with the active sites of IPU and Dex49A, the amino acid residues participating in subsites +2 and +3 are not conserved, and the glucose residues of isopanose bound to IPU completely differ in orientation from the corresponding glucose residues of isomaltose bound to Dex49A. The shape of the catalytic cleft characterized by the seventh coil of the beta-helix and a loop from domain N appears to be critical in determining the specificity of IPU for pullulan.

Selected figure(s)

Figure 5.
Fig. 5. Stereo view of the active sites of the IPU–isopanose (a) and Dex49A–isomaltose (b) complex forms. Isopanose and isomaltose are shown in red and blue, respectively. Residues conserved between IPU and Dex49A are shown in green, and residues that are unique to IPU or Dex49A are shown in orange. Two C^α backbones, loop N[A] and turn T[3] of coil 7, are shown in pink and yellow, respectively.

Figure 6.
Fig. 6. Stereo view showing the superposition of the C^α backbones between IPU (yellow) and endoPG II (gray). Three catalytic residues of IPU (Asp353, Asp372 and Asp373) and endoPG II (Asp180, Asp201 and Asp202) are shown in red and blue, respectively. Ser91 of endoPG II is also shown in blue. The numbering of the coils of the β-helix is as described in Fig. 2b. T[1] turns of coils 2 and 3 are indicated.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2008, 376, 210-220) copyright 2008.

Figures were selected by an automated process.