Structural basis of exo-β-mannanase activity in the GH2 family
| Autor: | Mário T. Murakami, Fabio C. Gozzo, Camila R. Santos, Letícia Maria Zanphorlin, Plínio Salmazo Vieira, Mariane Noronha Domingues, Renan A. S. Pirolla, Rodrigo V. Honorato, Paulo S. Oliveira, Mariana Abrahão Bueno de Morais, Flavio Henrique Moreira Souza |
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| Rok vydání: | 2018 |
| Předmět: |
Models
Molecular 0301 basic medicine Xanthomonas Protein Conformation Stereochemistry Mannose Crystallography X-Ray Biochemistry Substrate Specificity Enzyme catalysis Mannans 03 medical and health sciences chemistry.chemical_compound Protein structure Bacterial Proteins X-Ray Diffraction Catalytic Domain Scattering Small Angle Mannobiose Glycoside hydrolase Amino Acid Sequence Mode of action Molecular Biology chemistry.chemical_classification 030102 biochemistry & molecular biology biology Hydrolysis beta-Mannosidase Active site Cell Biology Kinetics 030104 developmental biology Enzyme chemistry Enzymology biology.protein Sequence Alignment |
| Zdroj: | Journal of Biological Chemistry. 293:13636-13649 |
| ISSN: | 0021-9258 |
| Popis: | The classical microbial strategy for depolymerization of β-mannan polysaccharides involves the synergistic action of at least two enzymes, endo-1,4-β-mannanases and β-mannosidases. In this work, we describe the first exo-β-mannanase from the GH2 family, isolated from Xanthomonas axonopodis pv. citri (XacMan2A), which can efficiently hydrolyze both manno-oligosaccharides and β-mannan into mannose. It represents a valuable process simplification in the microbial carbon uptake that could be of potential industrial interest. Biochemical assays revealed a progressive increase in the hydrolysis rates from mannobiose to mannohexaose, which distinguishes XacMan2A from the known GH2 β-mannosidases. Crystallographic analysis indicates that the active-site topology of XacMan2A underwent profound structural changes at the positive-subsite region, by the removal of the physical barrier canonically observed in GH2 β-mannosidases, generating a more open and accessible active site with additional productive positive subsites. Besides that, XacMan2A contains two residue substitutions in relation to typical GH2 β-mannosidases, Gly(439) and Gly(556), which alter the active site volume and are essential to its mode of action. Interestingly, the only other mechanistically characterized mannose-releasing exo-β-mannanase so far is from the GH5 family, and its mode of action was attributed to the emergence of a blocking loop at the negative-subsite region of a cleft-like active site, whereas in XacMan2A, the same activity can be explained by the removal of steric barriers at the positive-subsite region in an originally pocket-like active site. Therefore, the GH2 exo-β-mannanase represents a distinct molecular route to this rare activity, expanding our knowledge about functional convergence mechanisms in carbohydrate-active enzymes. |
| Databáze: | OpenAIRE |
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