Singh, Ratna; Weikert, Tobias; Basa, Sven; Moerschbacher, Bruno: Structural and biochemical insight into mode of action and subsite specificity of a chitosan degrading enzyme from Bacillus spec. MN
Inhalt
- Hydrolysis of fully deacetylated chitosan oligomers (GlcNn).
- Figure 1 Chitosan oligomer and polymer degradation with CSN-MN.
- Figure 2 Enzyme kinetics and MS spectrum for product pattern.
- Figure 3 Subsite specificity from MS2.
- Table 1 MS and MS2 analysis of products obtained from enzymatic cleavage of DA 30% chitosan polymer.
- Figure 4 Molecular modeling studies defining the property of the binding site.
- Figure 5 Stable hydrogen bonds between enzyme and substrate during simulation.
- Figure 6 Conformational changes occur at the binding site of the enzyme in the presence of substrate, and ensemble docking results.
- Figure 7 Extra subsite for binding GlcN subunit.
- Conclusions
- Figure 8 PCA analysis describing the states of the loops derived from substrate-free and -bound enzyme.
- Materials and Methods
- Bacterial strain, culture conditions, and enzyme purification.
- Oligomeric and polymeric chitosan substrates.
- Thin layer chromatography (TLC) of oligomeric products.
- Enzymatic hydrolysis of fully De-N-acetylated chitosan polymer.
- Enzymatic hydrolysis of partially acetylated chitosan polymers.
- Reducing end assay.
- MALDI-TOF MS analysis of chitosan oligomers.
- ESI MS analysis of chitosan oligomers.
- Molecular modelling.
- Electrostatic charge distribution and docking studies.
- Molecular dynamics simulations.
- Acknowledgements