Purification strategies of Scytalidium thermophilum xylanase

International Enzymes for Biocatalysis Workshop 3-5 June 2014, ISTANBUL

Purification strategies of Scytalidium thermophilum xylanase

1

Department of Food Engineering, Karamanoglu Mehmetbey University, Karaman, Turkey

2

Department of Chemistry, Karamanoglu Mehmetbey University, Karaman, Turkey

3

Department of Biology, Karamanoglu Mehmetbey University, Karaman, Turkey

Keywords: Xylanase, Scytalidium thermophilum, purification, column chromatography

Abstract

Xylanase (E.C.3.2.1.8) is an enzyme which hydrolyses β-1,4 bonds in xylan backbone which consists of D-xylose molecules. Based on its function, xylanase is used in many industries. For animal feed, paper production and textile industry partially purified xylanase can be acceptable, but for bakery and beverage industry, chemical or pharmaceutical applications, high purity is generally required.

In this study, xylanase was produced from a thermophilic fungus, Scytalidium thermophilum and purification strategies were developed for industrial and analytical purposes. In addition, within the cost reduction approach, corn cobs are used as carbon source in the culture medium which are produced millions of tons as agricultural waste each year and have very low economical value.

Partial purification of xylanase was studied with ammonium sulphate precipitation (ASP), aqueous two phase system (ATPS) and ultrafiltration (UF). Precipitation with the usage of 50% (v/v) ammonium sulphate concentration gave the best results as 2.5 fold purification and

52% yield. TX-114 was employed at a concentration of 7% (v/v) for ATPS and 2.7 fold

purification with 79% yield was obtained. Three-step UF technique was practiced using membranes with different MWCO values; which were 100, 30 and 10 kDa, sequentially. As the result of this technique, 4.3 fold purification with 25% yield was attained. With these results, partially purified xylanase can be presented as an ingredient for industrial applications.

For analytical purification of enzyme, two-step column chromatography technique, including gel filtration and anion exchange, was set up. Success of analytical purification was confirmed with SDS-PAGE electrophoresis which shows that xylanase was purified to homogeneity. By using a two-step chromatographic technique, without any additional pre-purification steps, xylanase was purified 21.8 fold with 9.6% yield which offers analytical purity where necessary.

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