Xylanase from a soil isolate, Bacillus pumilus : gene isolation, enzyme production, purification, characterization and one-step separation by aqueous-two-phase system (original) (raw)
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PURIFICATION OF XYLANASE FROM BACILLUS PUMILUS AND IT’S CHARACTERIZATION
Abstract This study is aimed at purification and characterization of xylanase produced by thermophilic and alkalophilic Bacillus pumilus isolated from corncob decaying soil. Culture supernatant (120.6Umg-1) of B. pumilus the xylanase was purified by ammonium sulphate precipitation and Sephadex G 75 gel filtration. With different concentrations of (NH4)2SO4, maximum amount of xylanase was precipitated at 50% of (NH4)2SO4 saturation. This (NH4)2SO4 precipitated sample was dialysed against distilled water for 24h and the sample (824.72 Umg-1) was loaded to Sephadex G 75 column and eluted with 0.5M Tris buffer at the flow rate of 0.5mL/min. Eluted fractions which showed highest xylanase activity were pooled together (2250.13 Umg-1), separated by Sodium Dodecyl Sulphate polyacrylamide (SDS) gel electrophoresis. Purified xylanase showed 2250.13 Umg-1 specific activitiy and the purification fold was18.6. The specific activity of the initial crude xylanase was 120.62 Umg-1 with a recovery y...
Production and characterization of xylanases of a Bacillus strain isolated from soil
World Journal of Microbiology & Biotechnology, 2005
Xylanase production was performed by growing a Bacillus isolate on agricultural by-products, wheat straw, wheat bran, corn cobs and cotton bagasse. A maximum xylanase activity of 180 U/ml was obtained together with a cellulase activity of 0.03 U/ml on 4 (w/v) corn cobs. Electrophoretic analysis showed the presence of three endo-β-1, 4-xylanases having molecular weights of about 22, 23 and 40 kDa. Xylanolytic activity was stable up to 50 °C in the pH range of 4.5–10 and the highest activity was observed at 70 °C and pH 6.5.
Production of alkali tolerant cellulase free xylanase in high levels by Bacillus pumilus SV-205
International Journal of Biological Macromolecules, 2012
The fermentation conditions were optimized for hyper production of xylanase from Bacillus pumilus SV-205. The bacterium secretes high levels (7382.7 ± 1200 IU/mL) of cellulase-free xylanase using wheat bran led to 21.63 fold increase in activity. A combination of yeast extract and peptone stimulated highest xylanase production (2448.0 IU/mL) as compared to other combinations. The most important characteristic of the enzyme is its high pH stability (100%) over a broad pH range of 6-11 for 24 h. Thermostability studies revealed that enzyme retained 65% activity after an incubation of 2 h at 60 • C. The level of production is remarkable as compared to earlier reports.
Characterization of xylanase produced by Bacillus pumilus
Journal of the National Science Foundation of Sri Lanka
This study was aimed at purifying xylanase produced by Bacillus pumilus. The spent medium contained 27.9 UmL -1 xylanase activity and 1.5 mgmL -1 protein. The highest specific activity (33.7 Umg -1 protein) was achieved with 50 % (NH 4 ) 2 SO 4 saturation and the xylanase recovery was 94.8 %.
Purification of xylanase produced by Bacillus pumilus
Journal of the National Science Foundation of Sri Lanka, 2014
This study was aimed at purifying xylanase produced by Bacillus pumilus. The spent medium contained 27.9 UmL -1 xylanase activity and 1.5 mgmL -1 protein. The highest specific activity (33.7 Umg -1 protein) was achieved with 50 % (NH 4 ) 2 SO 4 saturation and the xylanase recovery was 94.8 %.
Applied Ecology and Environmental Sciences, 2021
Xylanases are extensively applied in paper and pulp industries as well as during preparation of baked products to improve their quality. Additionally, it is also used in coffee, oil and starch industries in order to increase their nutritional values. Soil samples were collected near saw mills in various localities of Bangalore urban to isolate organisms for the production of xylanase using solid state fermentation. Six organisms were isolated using selective media based on their morphological characters. Among them one organism showed maximum production of xylanase enzyme identified as Bacillus sp based on their biochemical test and 16s RNA sequencing. Solid substrate fermentation was carried out using various agro wastes such as sugar cane bagasse, saw dust, paddy husk, wheat straw and orange peel powder. Sugarcane bagasse showed maximum production of enzyme compared to other substrates with different physical parameters such as pH 8, temperature at 35 °C after 72 hrs of incubation. Trace element such as Mg ++ enhances the production of enzyme more than 22 % compared with other metal ions like Ca ++ , Mn ++ and Fe ++. After production, enzyme purified by using three step methods such ammonium sulphate precipitation, dialysis, ion exchange and gel filtration. Fold purification was increased up to 12 fold, yield 36 % and molecular weight of enzyme was 62 KDa determined using SDS PAGE.
MIcrobiology Indonesia, 2009
Bacillus sp. AQ-1 was isolated from household aquarium sediment. The isolate produced extracellular xylanolytic enzymes on xylan containing agar medium. Based on morphological, and physiological analysis, the isolate was identified as Bacillus sp. AQ1. The effect of temperature and pH on isolate growth and xylanase production were observed. The best condition observed for the enzyme production in Luria Broth supplemented with 0.5% oat spelt xylan medium was at 40 °C pH 7. The maximum enzyme production was 0.23 U mL -1 after 20 h of fermentation. Two different medium compositions (A and B) were examined for xylanase production. The maximum growth of the isolate and the xylanase production was better in A medium. Replacing oat spelt xylan in medium A with fruitless oil palm bunch in the medium caused the growth slightly slower than that of in the original formula. However, the xylanase production was 3 times higher in fruitless oil palm bunch medium. Optimum activity of the crude enzyme was observed at 60 °C and pH 7. Each ml of the crude enzyme contained 55.21 U xylanase, 8.12 U amylase and 0.50 U carboxymethylcellulase.
Isolation and identification of local Bacillus isolates for xylanase biosynthesis
Iranian journal of microbiology, 2014
Bacillus species are attractive industrial organisms due to their rapid growth rates leading to a short fermentation cycle and for their capacity to secrete important enzymes and proteins such as xylanase into the extracellular medium. Considering the industrial importance of xylanase, in this current study, Bacillus spp. were isolated from different soils and were screened for their xylanase production. Bacillus isolates used in this study were obtained from a national screening program carried out during 2006-2007 in which soil samples that covered areas throughout the interior of Syria were collected. The prepared inoculum from each of Bacillus isolates was aliquoted onto xylan agar plates, incubated at 30°C for 72 h and screened for xylanase synthesis. Xylanolytic isolates were selected depending on the clear zones of xylan hydrolysis. Fifteen isolates having the highest clearing zone were determined and grown in a solid state fermentation. Of the 15 isolates, three bacilli name...
2015
The present study was aimed at isolation and characterization of a new xylan degrading strain of Bacillus from soil for production of xylanase. Different soil samples from Punjab state were used to isolate xylan degrading strains. We isolated a highly active xylanolytic strain and performed genetic characterisation using 16S r-RNA analysis. DNA sequencing and phylogenetic analysis using MEGA4 software showed our strain to be a novel strain (accession number JQ916900) which was named Bacillus lpuarvinder st. lpu002. Biochemical characterization of the enzyme showed optimum activity at 60 o C and pH 8.6. The enzyme was stable at 70 o C for up to 6 hours. The xylanase activity seems to be alleviated by divalent Ca 2+ , Mn 2+ and Mg 2+ ions. The best enzyme activity was obtained when the cells were grown in Bushnell-Haas medium supplemented with 0.5% xylan and beef extract. The process optimization was done using solid state fermentation (SSF) using wheat straw as substrate. The enzyme ...