Enzymatic characterization and molecular modeling of an evolutionarily interesting fungal β-N-acetylhexosaminidase
Language English Country Great Britain, England Media print-electronic
Document type Comparative Study, Journal Article, Research Support, Non-U.S. Gov't
- MeSH
- beta-N-Acetylhexosaminidases chemistry genetics isolation & purification metabolism MeSH
- Fungal Proteins chemistry genetics isolation & purification metabolism MeSH
- Glycosylation MeSH
- Catalytic Domain MeSH
- Kinetics MeSH
- Hydrogen-Ion Concentration MeSH
- Conserved Sequence MeSH
- Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase metabolism MeSH
- Models, Molecular * MeSH
- Molecular Sequence Data MeSH
- Penicillium enzymology genetics MeSH
- Enzyme Precursors chemistry genetics isolation & purification metabolism MeSH
- Protein Structure, Secondary MeSH
- Amino Acid Sequence MeSH
- Sequence Homology, Amino Acid MeSH
- Sequence Alignment MeSH
- Molecular Dynamics Simulation MeSH
- Enzyme Stability MeSH
- Substrate Specificity MeSH
- Temperature MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Comparative Study MeSH
- Names of Substances
- beta-N-Acetylhexosaminidases MeSH
- Fungal Proteins MeSH
- Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase MeSH
- Enzyme Precursors MeSH
Fungal β-N-acetylhexosaminidases are inducible extracellular enzymes with many biotechnological applications. The enzyme from Penicillium oxalicum has unique enzymatic properties despite its close evolutionary relationship with other fungal hexosaminidases. It has high GalNAcase activity, tolerates substrates with the modified N-acyl group better and has some other unusual catalytic properties. In order to understand these features, we performed isolation, biochemical and enzymological characterization, molecular cloning and molecular modelling. The native enzyme is composed of two catalytic units (65 kDa each) and two propeptides (15 kDa each), yielding a molecular weight of 160 kDa. Enzyme deglycosylated by endoglycosidase H had comparable activity, but reduced stability. We have cloned and sequenced the gene coding for the entire hexosaminidase from P. oxalicum. Sufficient sequence identity of this hexosaminidase with the structurally solved enzymes from bacteria and humans with complete conservation of all catalytic residues allowed us to construct a molecular model of the enzyme. Results from molecular dynamics simulations and substrate docking supported the experimental kinetic and substrate specificity data and provided a molecular explanation for why the hexosaminidase from P. oxalicum is unique among the family of fungal hexosaminidases.
References provided by Crossref.org
GENBANK
EU189026
