Protein engineering study of beta-mannosidase to set up a potential chemically efficient biocatalyst
| Authors | |
|---|---|
| Year of publication | 2014 |
| Type | Article in Periodical |
| Magazine / Source | Glycobiology |
| MU Faculty or unit | |
| Citation | |
| web | http://glycob.oxfordjournals.org/content/24/12/1301.long |
| Doi | http://dx.doi.org/10.1093/glycob/cwu074 |
| Field | Biochemistry |
| Keywords | Docking beta-mannosidase molecular dynamics mutagenesis protein engineering |
| Description | This study is focused on the analysis and mutagenesis of beta-mannosidase from Bacteroides thetaiotaomicron with the aim of broadening its substrate specificity to 2-acetamido-2-deoxy-beta-d-mannopyranosyl (beta-ManNAc) derivatives. Various conformations (4C1, 4H5, and 1S5) of native and modified ligands were docked to the binding site of the protein to determine the most suitable conformation of sugars for further hydrolysis. Key amino acid residues were mutated in silico focusing on stabilizing the acetamido group of beta-ManNAc as well as forming the oxazoline intermediate needed for hydrolysis. The results of large set of 5 ns molecular dynamic simulations showed that the majority of the active site residues are involved in substrate interaction and do not exhibit a higher flexibility except for Asn178. Mutations of Asn178 to alanine and Asp199 to serine could lead to a stabilisation of the acetamido group in the binding site. So far, in vitro mutagenesis and the screen of a large variety of biological sources were unable to extend beta-mannosidase's activity to include beta-ManNAc derivatives. |
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