Heterologous Expression of a Thermostable Chitinase from <i>Myxococcus xanthus</i> and Its Application for High Yield Production of Glucosamine from Shrimp Shell
Glucosamine (GlcN) is a widely used food supplement. Hence, enormous attention has been concerned with enzymatic production of GlcN owing to its advantage over a chemical approach. In this study, a previously unstudied chitinase gene (MxChi) in the genome of <i>Myxococcus xanthus</i> was...
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| Auteurs principaux: | , , , , , , , , |
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| Format: | article |
| Langue: | EN |
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MDPI AG
2021
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| Accès en ligne: | https://doaj.org/article/c86b135d1f694c768717bbcf9bb5c2c1 |
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| Résumé: | Glucosamine (GlcN) is a widely used food supplement. Hence, enormous attention has been concerned with enzymatic production of GlcN owing to its advantage over a chemical approach. In this study, a previously unstudied chitinase gene (MxChi) in the genome of <i>Myxococcus xanthus</i> was cloned, expressed in recombinant soluble form and purified to homogeneity. TLC-, UPLC-, and microplate-reader- based activity tests confirmed MxChi hydrolyzes colloidal chitin to chitobiose as sole product. The optimal catalytic pH and temperature of MxChi was identified as 7.0 and 55 °C, respectively. MxChi exhibited 80% activity after 72 h incubation at 37 °C. The site-directed mutagenesis revealed that the amino acids D323A, D325A, and E327A of MxChi were in the DXDXE catalytic motif of GH18. When coupled with β-N-acetylhexosaminidase (SnHex) and deacetylase (CmCBDA), the enzyme allowed one-pot extraction of GlcN from colloidal chitin and shrimp shell. The optimal condition was 37 °C, pH 8.0, and 1/3/16.5 (MxChi/SnHex/CmCBDA), conducted by orthogonal design for the enzymatic cascades. Under this condition, the yield of GlcN was 26.33 mg from 400 mg shrimp shell. Facile recombinant in <i>E. coli</i>, robust thermostability and pure product herein makes newly discovered chitinase a valuable candidate for the green recycling of chitin rich waste. |
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