Production Of High Activity Xylanases By Two Streptomyces Strains And The Enzyme Properties

Microbial xylanases have attracted considerable research interest in recent years because their potential application in the food, animal feed, paper and pulp industries.In the paper, xylanase producing stains were isolated from soil by plate screening with xylan as the sole carbon source, and among them a high xylanase producing strain was selected by experiments in shake flasks. By observation of its colonies, mycelia, spores, and by comparison its physicochemical properties with typical stain's (Streptomyces cirratus ATCC14699), it was identified as Streptomyces cirratus D-10,Eleven xylanase producing improved stains were screened by penicillin containing plate after spores of Streptomyces cirratus D-10 were treated with UV-irradiation and ethyl-methanesulfonate.The effects of different factors on xylanase production by Streptomyces cirratus D-10 were studied in shake flask condition. The best initial pH value of growth medium for xylanase production was at pH6.0. Corn cob xylan (water insoluble) and Tryptone were the best C and N sources, respectively. The enzyme activity was increased to about 2.5 times by addition of 0.5% Tween 80 in the medium. The highest xylanase activity was up to 623u/mL.A xylanase was separated and purified from the culture filtrate of Streptomyces cirratus D-10 by ammonium sulphate precipitation and Q-sephrose FF ion exchange chromatography. The molecular weight of the enzyme was estimated to be 22.0 KDa by SDS-PAGE. After a two-step treatment, ten-fold purification was obtained with 24% enzyme activity recovery.The purified xylanase shows maximal activity at pH7.50. It remains 90% activity at pH ranges between pH 3.10 and pH 9.80 under 45 . The enzyme exhibits maximal activity at 65 and shows desirable thermal stability under 50 .At 1mM concentration, EDTA and SDS have little influence on the xylanse activity. The inhibit effects of different metal ions on the enzyme activity are as follows: Ag+ > Pb2+ , Mn2+ >Cu2+, Co2+ > Ba2+, Mg2+, Ca2+ , Na+, K+, Zn2+. The only metal ion which can improve the enzyme activity is Fe2+.The enzyme shows higher activity on oatspelts xylan (water insoluble), beechwood xylan and birchwood xylan; shows lower activity on oatspelts xylan (water soluble) and least activity on self-made corn cob xylan and cotton seed husk xylan.The effects of different factors on xylanase production by Streptomyces olivaceoviridis E-86 were also studied in shake flask condition. The best initial pH value of growth medium for xylanase production was at pH6.0. Corn cob xylan and beef peptone were the best C source and N source, respectively. The enzyme activity was increased to about 2 times by addition of 1.5%(v/v) Tween 80 inthe medium. The highest xylanase activity obtained was 1653 u/ml which was also the highest ever reported from Streptomyces sp.The highest xylanase activities in a 5-1 bioreactor and a 200-1 reactor were 540 u/ml and 625 u/ml, respectively, by Streptomyces olivaceoviridis E-86. No report was seen about xylanase production by Streptomyces sp. in a 200-1 and larger reactor.The native-PAGE and zymogram showed that in native condition the molecular weight of active enzyme protein in the culture filtrate of Streptomyces olivaceoviridis E-86 was estimated to be about 750 KDa which was also firstly reported.