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Substrate recognition of N,N'-diacetylchitobiose deacetylase from Pyrococcus horikoshii.
- Source :
-
Journal of structural biology [J Struct Biol] 2016 Sep; Vol. 195 (3), pp. 286-293. Date of Electronic Publication: 2016 Jul 22. - Publication Year :
- 2016
-
Abstract
- Enzymes of carbohydrate esterase (CE) family 14 catalyze hydrolysis of N-acetyl groups at the non-reducing end of the N-acetylglucosamine (GlcNAc) residue of chitooligosaccharides or related compounds. N,N'-diacetylchitobiose deacetylase (Dac) belongs to the CE-14 family and plays a role in the chitinolytic pathway in archaea by deacetylating N,N'-diacetylchitobiose (GlcNAc2), which is the end product of chitinase. In this study, we revealed the structural basis of reaction specificity in CE-14 deacetylases by solving a crystal structure of Dac from Pyrococcus horikoshii (Ph-Dac) in complex with a novel reaction intermediate analog. We developed 2-deoxy-2-methylphosphoramido-d-glucose (MPG) as the analog of the tetrahedral oxyanion intermediate of the monosaccharide substrate GlcNAc. The crystal structure of Ph-Dac in complex with MPG demonstrated that Arg92, Asp115, and His152 side chains interact with hydroxyl groups of the glucose moiety of the non-reducing-end GlcNAc residue. The amino acid residues responsible for recognition of the MPG glucose moiety are spatially conserved in other CE-14 deacetylases. Molecular dynamics simulation of the structure of the Ph-Dac-GlcNAc2 complex indicated that the reducing GlcNAc residue is placed in a large intermolecular cleft and is not involved with specific interactions with the enzyme. This observation was consistent with results indicating that Ph-Dac displayed similar kinetic parameters for both GlcNAc and GlcNAc2. This study provides the structural basis of reaction-site specificity of Dac and related CE-14 enzymes.<br /> (Copyright © 2016 Elsevier Inc. All rights reserved.)
- Subjects :
- Amino Acid Sequence
Catalytic Domain
Chitin analogs & derivatives
Chitosan
Crystallography, X-Ray
Hydrogen Bonding
Kinetics
Molecular Dynamics Simulation
Oligosaccharides
Phosphates chemistry
Substrate Specificity
Archaeal Proteins chemistry
Disaccharides chemistry
Pyrococcus horikoshii enzymology
Subjects
Details
- Language :
- English
- ISSN :
- 1095-8657
- Volume :
- 195
- Issue :
- 3
- Database :
- MEDLINE
- Journal :
- Journal of structural biology
- Publication Type :
- Academic Journal
- Accession number :
- 27456364
- Full Text :
- https://doi.org/10.1016/j.jsb.2016.07.015