HAYATI Journal of Biosciences, September 2007, p 115-118 Vol. 14, No. 3 ISSN: 1978-3019 SHORT COMMUNICATION Characterization of Xylanase from Streptomyces spp. Strain C1-3 ANJA MERYANDINI Department of Biology, FMIPA, Bogor Agricultural University, Darmaga Campus, Bogor 16680, Indonesia Phone/Fax: +62-251-622833, E-mail: ameryandini@yahoo.com Received April 6, 2006/Accepted September 7, 2007 Xylan is the major constituent of hemi cellulose. Several enzymes are needed to hydrolyse xylan completely, includ- ing xylanase. Currently, there is an increasing use of this enzyme. This study was carried out to characterize the xylanase from Streptomyces spp. strain C1-3. Results showed that the xylanase displayed its highest activity at pH 3 and 90 o C and was stable up to 10 hours at this conditions. Its activity increased after the addition of Cu 2+ , Fe 2+ , and Co 2+ under concentration of 1 and 5 mM, respectively. The activity however, decreased after the addition of Mg 2+ , Ca 2+ at 1 mM and Zn 2+ at 5 mM. After a test with five kinds of xylan (i.e. from Birchwood, Beechwood, Arabinoxylan, Oat spelt and CMC), the xylanase of Streptomyces spp. C1-3 showed its preferences to Birchwood- and Arabino-xylan. The results showed that the xylanase of Streptomyces spp. C1-3 was characterized as a thermostable acid xylanase. Key words: xylanase, Streptomyces, stability, CMCase ___________________________________________________________________________ Hemicellulose is the second most abundant hetero- polysaccharides after cellulose and xylan is the major constituent of it. Xylan has a backbone chain of 1,4 linked β-D xylopiranose units. The backbone consists of O-acetyl, α-L-arabinocyl, 4-metilglucuronic acid. An enzyme system is needed to hydrolyze xylan completely. This enzyme system consist of endo-β-1,4 xylanase, β-xilosidase, galactosidase, α-arabinofuranosidase, α-D-glucuronidase, and acetyl xylan esterase (Sunna et al. 1997; Subramaniyan & Prema 2002). Currently, xylanase application showed highly increase due to its necessity of the products produced from dissolving pulp such as rayon, cellophane, and chemicals. Xylanase is also used in bioconversion of lignocellulotic material, agro- waste products, clarifying juice and feedstock. As xylan hydrolysis is a main factor in all processes above (Beg et al. 2001; Subramaniyan & Prema 2002), hence the use of xylanase for all those purposes should be characterized. Xylanase is produced by fungi (Lin et al. 1999; Saha 2002; Ryan et al. 2003), bacteria (Beg et al. 2001), and protozoa (Devillard et al. 2003). One of the potential bacteria group that produced xylanase is Actinomycetes, especially Streptomyces (Ruiz-arribas et al. 1995; Georis et al. 2000; Kaneko et al. 2000; Wang et al. 2003; Kansoh & Nagieb 2004). The aim of this research was to characterize Streptomyces spp. C1-3 xylanase isolated from Cicurug Sukabumi soil sample. It was one of several Streptomyces collections we had in Dept. of Biology. The Streptomyces spp. C1-3 isolate was rejuvenated in YM agar-agar media (0.4% yeast extract, 1% malt extract, 1.5% glucose, and 1.5% agar-agar). The isolate was then grown in a xylan media (1% yeast extract, 10.3% sucrose, 0.5% Birchwood Xylan, 1.5% agar-agar) and was incubated at 30 o C for seven days. Two cockbors (diameter = 2 cm) of the isolate grown in the xylan media was subsequenthly inoculated to 100 ml xylan media in 500 ml Erlenmeyer. They were incubated in 140 rpm agitation at 30 o C for 10 days. The culture was centrifuged every day (5 minutes) at 10,000 x g to obtain the xylanase crude extract. The extract activity was measured by using DNS (Dinitrosalisilic Acid) method of Miller (1959) with xylosa as the standard. The reducing sugar of the reference samples (substrate solutions incubated without enzyme and a diluted enzyme solution in a buffer) was deduced from the values of the test samples. The yielded reducing-sugar substance was assessed by spectrophotometer (λ = 540 nm). One unit xylanase activity was defined as the amount of enzyme producing 1 μmol xylosa per minute. Protein concentration (mg/ml) was defined by using Bradford method (1976) and Bovine serum albumin (BSA) was used as the standard. Characterization of the crude extract enzyme were carried out to determine the optimum temperature and pH, enzyme stability, and the influence of bivalent cations. All data in this study were from duplo trials. The assessment of pH was carried out within a pH range of 3.0-9.0 with 0.5 interval. The determination of optimum temperature was performed from 30 up to 90 o C with 10 o C interval. The stability of xylanase crude extract was tested by incubating the extract without substrate in two different temperatures the first at its optimum temperature and the second at 3 o C of storage temperature. To observe the influence of cations on the enzyme activity, six cations (i.e. Ca 2+ , Zn 2+ , Cu 2+ , Mg 2+ , Fe 2+ , and Co 2+ ) derived from CaCl 2 , ZnCl 2 , CuCl 2 , MgCl 2 , FeSO 4 ·7H 2 O, and CoCl 2 , were added separately to a final concentration of 1 and 5 mM. Determining the enzyme activity in several kinds of xylan substrates was defined by assessing the extract activity in Birchwood Xylan, Oatspelt Xylan, Wheat Arabinoxylan, Beechwood Xylan, and Carboxy Methyl Cellulose (CMC).