Purification and characterization of an acid-stable and organic solvent-tolerant xylanase from Aspergillus awamori VTCC-F312

Abstract

An acid-stable and organic solvent-tolerant extracellular xylanase was isolated and purified from Aspergillus awamori VTCC-F312 and its properties were investigated. The xylanase was purified by Sephadex G-100 gel filtration chromatography and DEAE-Sephadex A-50 ion exchange chromatography to homogeneity. The enzyme had a molecular mass of 32 kDa and a specific activity of 217 U/mg protein, with optimum temperature of 50-55°C and optimum pH of 5. This enzyme was stable at up to 45°C and no loss of enzyme activity was observed after incubation for 6 h at 40°C. The xylanase was stable within the pH range of 4-8 with no loss of enzyme activity after incubation at 30°C for 1-4 h, even the enzyme activity was found to increase by 60% after incubation at pH 4 for 4 h. The Km and Vmax values were 12.6 mg oat spelt xylan per ml and 1000 U/mg protein, respectively. Dithiothreitol, β-mercaptoethanol, and EDTA were found to increase the xylanase activity by 19%, 46%, and 138%, respectively. Except for Fe 3+ , the presence of 5 mM tested metal ions increased the enzyme activity by 18-52%. The enzyme showed high resistance to organic solvents and retained 63-86% and 44-61% of its activity in the presence of tested organic solvents at 30% and 80% (v/v), respectively. Triton X-100 and Tween 80 activated the xylanase to 128% and 116% of its activity, whereas SDS at the concentration of 5% completely inhibited the enzyme. These results suggested that the xylanase from A. awamori VTCC-F312 could potentially be used as a feed additive for monogastric animals.KEYWORDS: detergent-resistant, no disulphide bond, non-metal enzyme, free of cellulase and mannanase activity, corn cob, natural substrate

Aspergillus sydowii: Genome Analysis and Characterization of Two Heterologous Expressed, Non-redundant Xylanases

“…Reducing agents, like DTT (for Fsh102) or β-Mercaptoethanol (for SBS 45 xylanases), do not affect the enzymes as no disulfide bonds are present. By comparing the enzymes to a xylanase from A. niger DSM 1957 (Do et al, 2009), which is stated to be resistant to methanol, ethanol, isopropanol, and acetone up to 30 (v/v)%, or to Aspergillus awamori VTCC-F312 xylanase with has a residual activity of 63-86% by 30 (v/v)% solvents (Do et al, 2012), the Fsh102 xylanases can be classified as moderate solvent stable, which is necessary for the use some pulp and paper processing like the organosolve process (Brosse et al, 2019).…”