Partial purification and characterization of an extracellular Chitin Deacetylase from Streptomyces griseoinacarnatus RB7AG isolated from Chilika lake

Abstract

An extracellular thermo-alkali tolerant chitin deacetylase (CDA) was obtained from Streptomyces griseoincarnatus RB7AG, isolated from Chilika lake sediments. The purification of the enzyme was carried out by ammonium sulfate precipitation, followed by chromatographic separation by diethylaminoethanol sepharose (DEAE sepharose) ion exchange resin. This purification resulted in a 29.42-fold increase in purity, achieving a specific activity of 3.09 U/mg with a yield of 40%. The molecular weight of the enzyme was 50 kDa. The enzyme exhibited optimal activity at a temperature of 40 °C and a pH of 8.0, displaying remarkable stability over a wide range of temperatures (4–25 °C) and pH levels (6.0–9.0). It demonstrated enhanced enzymatic activity when exposed to certain metal ions, particularly Cu2+, Mg2+, and Fe3+. The enzyme exhibited Km and Vmax values of 1.55 mg/mL and 20.9 µmol/min. This CDA exhibited activity towards glycol chitin and chitin oligomers with a degree of deacetylation greater than 75%. MALDI-TOF/MS (Matrix-assisted laser desorption ionization-time-of-flight mass spectrometry) data revealed that the isolated protein was found to be close relatives of the polysaccharide deacetylase family and have four conserved CE4 (Carbohydrate esterase 4) family. Based on the SDS PAGE (Sodium dodecyl-sulfate polyacrylamide gel electrophoresis), activity assays, generation of a higher degree of deacetylated product, and MALDI-TOF/MS data, it is proposed that the purified enzyme from Streptomyces griseoincarnatus RB7AG is an extracellular chitin deacetylase.