Purification and characterization of xylanase from endophytic fungus Alternaria alternata isolate PTRa9

Abstract

Microorganisms were the major sources of xylanase production due to there were important in agriculture and industrial. The aim of this research was to study the xylanase production from endophytic fungi. Totally 54 endophytes isolated from Corton oblongifolius Roxb., Palm and mangrove leaves were tested for xylanase activity. There were only 31 endophytes isolated produce clear zone around colony in selective xylan agar medium which indicated xylanase-producing in primary screening. Endophyte isolate PTRa9 yield the highest xylanase-production in secondary screening. The optimal conditions that PTRa9 produced highest xylanase activity when 2% rice bran and 0.1% (NH₄)₂SO₄ as carbon and grew in the medium supplemented with nitrogen source, respectively for 4 days. Xylanase purification was done by ammonium sulfate precipitation, ion exchange chromatography and gel filtration chromatography which resulted in an apparent increase in specific activity as 161.1 U/mg protein and 60.8 times of purity. Xylanase characterization includes thermal stability, pH stability, metal ions, kinetic analysis and molecular weight were studied and the result showed that at 40°C for 20 min, at pH range 3 to 11 for 60 min had relative xylanase activity of 131.27 and higher than 100 %, respectively. PTRa9 xylanase was strongly inhibited by Hg²⁺, and was moderately inhibited by Mg²⁺, Mn²⁺, Cu²⁺, Zn²⁺, Fe²⁺ and EDTA. For kinetic analysis, it had Km of 2.369 mg/ml and Vmax of 2.142 µl/min/mg protein. Molecular weight of Xylanase of PTRa9 was of 54.8 kDa.