Journal
ANALYTICAL CHEMISTRY
Volume 90, Issue 18, Pages 10654-10658Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acs.analchem.8b02729
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Funding
- NANO3BIO project
- EU 7FP Grant [613931]
- MINECO, Spain [BFU2016-77427-C2-1-R]
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Not only the degree of acetylation but also the pattern of acetylation of chitosans and chitooligosaccharides (COS) appear to be critical for their biological activities. Protein engineering may expand the toolbox of chitin deacetylases (CDAs) with defined specificities for the enzymatic production of partially deacetylated COS for biotech and biomedical applications. A high-throughput screening (HTS) assay for screening directed evolution libraries is reported. It is based on a fluorescence monitoring assay of the deacetylase activity on COS substrates after capturing the expressed enzyme variants fused to a chitin binding module with chitin-coated magnetic beads. The assay is applied to the screening of random libraries of a Vibrio cholera CDA for increased activity on longer COS substrates.
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