Iason Tsigos

Chitin deacetylases: new, versatile tools in biotechnology

Chitin deacetylases have been identified in several fungi and insects. They catalyse the hydrolysis of N-acetamido bonds of chitin, converting it to chitosan. Chitosans, which are produced by a harsh thermochemical procedure, have several applications in areas such as biomedicine, food ingredients, cosmetics and pharmaceuticals. The use of chitin deacetylases for the conversion of chitin to chitosan, in contrast to the presently used chemical procedure, offers the possibility of a controlled, non-degradable process, resulting in the production of novel, well-defined chitosan oligomers and polymers.

Purification and Characterization of Chitin Deacetylase from Colletotrichum lindemuthianum

Chitin deacetylase (EC 3.5.1.41), the enzyme that catalyzes the hydrolysis of acetamido groups of N-acetyl-D-glucosamine in chitin, has been purified to homogeneity from the culture filtrate of the fungus Colletotrichum lindemuthianum and further characterized. The enzyme is a glycoprotein, and its apparent molecular mass was determined to be 150 kDa. The glycosylation pattern of the enzyme is consistent with a mixture of N-linked glycans including oligomannosidic hybrid and/or complex type, and its carbohydrate content is approximately 67% by weight. Chitin deacetylase is active on several chitinous substrates and chitin derivatives, is not considerably inhibited by carboxylic acids, especially acetic acid, and exhibits a remarkable thermostability. The enzyme requires at least two N-acetyl-D-glucosamine residues (chitobiose) for catalysis. When glycol chitin (a water-soluble chitin derivative) was used as substrate, the optimum temperature for enzyme activity was determined to be 50°C, and the optimum pH was 8.5.

Stereospecificity of hydrogen transfer by the NAD+-linked alcohol dehydrogenase from the Antarctic psychrophile Moraxella sp. TAE123

Investigation of the stereochemistry of the hydride transfer in reactions catalyzed by the recently isolated NAD(+)-linked alcohol dehydrogenase from the Antarctic psychrophile Moraxella sp. TAE123 was accomplished by using 1H NMR spectroscopy of the deuterated coenzyme. It was found that this new psychrophilic enzyme is a type A dehydrogenase. Moraxella sp. ADH reduces stereospecifically 2-butanone to produce (S)-2-butanol.

Crystallization and preliminary X-ray diffraction studies of an alcohol dehydrogenase from the Antarctic psychrophile Moraxella sp. TAE123

An NAD(+)-dependent psychrophilic alcohol dehydrogenase (ADH) from the Antarctic psychrophile Moraxella sp. TAE123 has been purified to homogeneity. The enzyme consists of four identical subunits, each containing two Zn ions. Protein crystals suitable for X-ray diffraction were obtained under optimized salting-out crystallization conditions using ammonium sulfate as a precipitating agent. The crystals are hexagonal bipyramids and belong to space group P3(1)21 or P3(2)21, with unit-cell parameters a = 136.4, c = 210.7 A. They contain one protein homotetramer in the asymmetric unit. Diffraction data were collected to 2.2 A under cryogenic conditions using synchrotron radiation.