SSF production, purification and characterization of chitin deacetylase from Aspergillus flavus

Abstract

The production of an extracellular chitin deacetylase (CDA) produced by Aspergillus flavus under solid-substrate fermentation (SSF) using wheat bran as substrate was optimized using statistical methods. The CDA production in SSF increased 1.79-fold in comparison to the unoptimized basal level medium. It was purified to a final purity of 3.94-fold by ammonium sulphate precipitation, ion-exchange chromatography, and gel-permeation chromatography (GPC) consecutively and further characterized. The molecular mass of the enzyme was estimated to be about 28 kDa by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and GPC analysis. The optimum pH and temperature of the purified enzyme were pH 8.0 and 50 °C, respectively. Additionally, the effect of some cations and other chemical compounds on the CDA activity was studied. A marginal increase in enzyme activity was observed with metal ions mainly Mn²⁺ and Zn²⁺. No inhibition of the enzyme was observed by the end product, that is, acetate up to 70 mM concentration. The K_m and k_cat values of the enzyme were determined to be 9.45 mg mL⁻¹ and 26.72 s⁻¹ respectively, using colloidal chitin as substrate. Among various substrates tested, glycol chitin and colloidal chitin were deacetylated.

Keywords:

• Chitin deacetylase
• Aspergillus flavus
• SSF production
• statistical optimization
• enzyme purification
• enzyme characterization

Acknowledgements

The authors are thankful to Council for Scientific and Industrial Research (CSIR), New Delhi for providing the CSIR-SRF fellowship to Narayanan Karthik.

Disclosure statement

The authors report no declarations of interest.