Cloning and expression of a chitin deacetylase gene (CDA2) from Saccharomyces cerevisiae in Escherichia coli: Purification and characterization of the cobalt-dependent recombinant enzyme

The chitin deacetylase gene CDA2 from Saccharomyces cerevisiae has been cloned and expressed in Escherichia coli and the recombinant enzyme purified to homogeneity and further characterized. The enzyme exhibits an apparent molecular mass of 35 kDa. When glycol chitin is used as substrate the optimum temperature for enzyme activity is 50 °C and the pH optimum is 8.0. The enzyme requires at least two N-acetyl-d-glucosamine residues (chitobiose) for catalysis and is inhibited by acetate. The presence of CoCl2 proved to be essential for enzyme activity. Seven amino acids could be eliminated from the C-terminus without a significant loss of enzyme activity. However, truncation of 12 or more amino acids from the N-terminus or 27 amino acids from the C-terminus resulted in complete loss of enzyme activity.