PDBsum entry 2gac

Hydrolase

PDB id: 2gac

Contents

Protein chains

136 a.a. *

139 a.a. *

Waters ×246

* Residue conservation analysis

PDB id:

2gac

Name:

Hydrolase

Title:

T152c mutant glycosylasparaginase from flavobacterium meningosepticum

Structure:

Glycosylasparaginase. Chain: a, c. Synonym: glycoasparaginase, aspartylglycosylaminase, aspartylglucosaminidase. Engineered: yes. Mutation: yes. Glycosylasparaginase. Chain: b, d. Synonym: glycoasparaginase, aspartylglycosylaminase,

Source:

Elizabethkingia meningoseptica. Organism_taxid: 238. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Hetero-Dimer (from PDB file)

Resolution:

2.10Å R-factor: 0.233 R-free: 0.280

Authors:

H.-C.Guo,Q.Xu

Key ref:

H.C.Guo et al. (1998). Crystal structures of Flavobacterium glycosylasparaginase. An N-terminal nucleophile hydrolase activated by intramolecular proteolysis. J Biol Chem, 273, 20205-20212. PubMed id: 9685368 DOI: 10.1074/jbc.273.32.20205

Date:

29-May-98 Release date: 08-Jun-99

Protein chains

Q47898  (ASPG_ELIMR) -  N(4)-(Beta-N-acetylglucosaminyl)-L-asparaginase from Elizabethkingia miricola

Seq: 340 a.a.

Struc: 136 a.a.

Protein chains

Q47898  (ASPG_ELIMR) -  N(4)-(Beta-N-acetylglucosaminyl)-L-asparaginase from Elizabethkingia miricola

Seq: 340 a.a.

Struc: 139 a.a.*

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

Enzyme reactions

• Enzyme class: Chains A, B, C, D: E.C.3.5.1.26 - N(4)-(beta-N-acetylglucosaminyl)-L-asparaginase.
• Reaction: N₄-(beta-N-acetyl-D-glucosaminyl)-L-asparagine + H2O = N-acetyl-beta-D- glucosaminylamine + L-aspartate + H⁺

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1074/jbc.273.32.20205

J Biol Chem 273:20205-20212 (1998)

PubMed id: 9685368

Crystal structures of Flavobacterium glycosylasparaginase. An N-terminal nucleophile hydrolase activated by intramolecular proteolysis.

H.C.Guo, Q.Xu, D.Buckley, C.Guan.

ABSTRACT

Glycosylasparaginase (GA) is a member of a novel family of N-terminal nucleophile hydrolases that catalytically use an N-terminal residue as both a polarizing base and a nucleophile. These enzymes are activated from a single chain precursor by intramolecular autoproteolysis to yield the N-terminal nucleophile. A deficiency of GA results in the human genetic disorder known as aspartylglycosaminuria. In this study, we report the crystal structure of recombinant GA from Flavobacterium meningosepticum. Similar to the human structure, the bacterial GA forms an alphabetabetaalpha sandwich. However, some significant differences are observed between the Flavobacterium and human structures. The active site of Flavobacterium glycosylasparaginase is in an open conformation when compared with the human structure. We also describe the structure of a mutant wherein the N-terminal nucleophile Thr152 is substituted by a cysteine. In the bacterial GA crystals, we observe a heterotetrameric structure similar to that found in the human structure, as well as that observed in solution for eukaryotic glycosylasparaginases. The results confirm the suitability of the bacterial enzyme as a model to study the consequences of mutations in aspartylglycosaminuria patients. They also suggest that further studies are necessary to understand the detail mechanism of this enzyme. The presence of the heterotetrameric structure in the crystals is significant because dimerization of precursors has been suggested in the human enzyme to be a prerequisite to trigger autoproteolysis.

Selected figure(s)

Figure 1.
Fig. 1. The structure of glycosylasparaginase from F. meningosepticum. a, stereo ribbon representation of the Flavobacterium GA structure. One heterodimer is shown with a- (red) and b-subunits (green). The active site is at top center of the structure toward the viewer and around the N-terminal end of the b-subunit (green) (labeled Nb in light blue). b, stereo diagram of C traces of Flavobacterium GA. c, stereo diagram of C traces of Flavobacterium (dark blue) and human (gray) GA. Superimposition is based on all common main chain atoms (excluding insertions/deletions). The residues labeled are in Flavobacterium sequence number.

Figure 4.
Fig. 4. Stereo view of the active site of glycosylasparaginase. a, stereo view of superimposition of the active sites between Flavobacterium (shown according to atom type: yellow for carbons, blue for nitrogens, and red for oxygens) and human (shown in gray) GA. Also shown is aspartate in the human enzyme/product structure (13). Dashed lines correspond to the hydrogen bonds described in the human structure. b, the same stereo view of active site in the T152C mutant. The color scheme is the same as the wild type in (a), except that sulfur of the thiol group is shown in green.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (1998, 273, 20205-20212) copyright 1998.

Figures were selected by an automated process.