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Cloning and heterologous expression of cellulose free thermostable xylanase from Bacillus brevis.

Goswami GK, Krishnamohan M, Nain V, Aggarwal C, Ramesh B - Springerplus (2014)

Bottom Line: The enzyme did not show any cellulose activity and was active under wide range of temperature (40°C to 80°C) and pH (4 to 9).The enzyme showed considerable thermo stability and regained over 90% of activity, when returned to 55°C after boiling for 5 min.These physiochemical properties of B. brevis xylanse show high potential of its applications in paper and pulp industry.

View Article: PubMed Central - PubMed

Affiliation: Amity Institute of Biotechnology, Amity University Rajasthan, Jaipur, 302001 India.

ABSTRACT
Xylanase gene isolated from Bacillus brevis was expressed in E. coli BL21. Sequencing of the gene (Gen Bank accession number: HQ179986) showed that it belongs to family 11 xylanases. The recombinant xylanase was predominantly secreted to culture medium and showed mesophilic nature (optimum activity at 55°C and pH 7.0). The cell free culture medium exhibited 30 IU/ml xylanse activity. The enzyme did not show any cellulose activity and was active under wide range of temperature (40°C to 80°C) and pH (4 to 9). The enzyme showed considerable thermo stability and regained over 90% of activity, when returned to 55°C after boiling for 5 min. These physiochemical properties of B. brevis xylanse show high potential of its applications in paper and pulp industry.

No MeSH data available.


Effect of temperature and pH on the xylanase activity. (A). Effect of temperature on enzyme activity (in IU) of B. brevis xylanase. (B). Effect of different pH levels on B. Brevis xylanase relative activity.
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Fig4: Effect of temperature and pH on the xylanase activity. (A). Effect of temperature on enzyme activity (in IU) of B. brevis xylanase. (B). Effect of different pH levels on B. Brevis xylanase relative activity.

Mentions: SDS-PAGE of IPTG induced Zymogram was developed that showed xylanase activity on the gel by clearing zone development on staining with Congo Red dye. Xylanase production was found more (approximately 1.5 times higher) in cloned expression host as compared to B. brevis (Figure 3). The optimization of temperature of both cloned and native (B. brevis) xylanases show mesophilic nature of B. brevis xylanase with maximum activity at 55°C Figure 4A. Similarly the optimum pH of the B. brevis xylanase was found at pH 7 (Figure 4B). After finding optimum temperature and pH, effect of high temperature on the enzyme activity was measured. Although enzyme showed considerable loss of enzyme activity at higher temperature and negligible activity was observed above 90°C, the enzyme regained more than 90% activity after boiling for 5 min and subsequent cooling at 37°C for 45 minutes. The enzyme was also evaluated for any cellulose activity and as expected, no detectable cellulase activity was observed.Figure 3


Cloning and heterologous expression of cellulose free thermostable xylanase from Bacillus brevis.

Goswami GK, Krishnamohan M, Nain V, Aggarwal C, Ramesh B - Springerplus (2014)

Effect of temperature and pH on the xylanase activity. (A). Effect of temperature on enzyme activity (in IU) of B. brevis xylanase. (B). Effect of different pH levels on B. Brevis xylanase relative activity.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4320173&req=5

Fig4: Effect of temperature and pH on the xylanase activity. (A). Effect of temperature on enzyme activity (in IU) of B. brevis xylanase. (B). Effect of different pH levels on B. Brevis xylanase relative activity.
Mentions: SDS-PAGE of IPTG induced Zymogram was developed that showed xylanase activity on the gel by clearing zone development on staining with Congo Red dye. Xylanase production was found more (approximately 1.5 times higher) in cloned expression host as compared to B. brevis (Figure 3). The optimization of temperature of both cloned and native (B. brevis) xylanases show mesophilic nature of B. brevis xylanase with maximum activity at 55°C Figure 4A. Similarly the optimum pH of the B. brevis xylanase was found at pH 7 (Figure 4B). After finding optimum temperature and pH, effect of high temperature on the enzyme activity was measured. Although enzyme showed considerable loss of enzyme activity at higher temperature and negligible activity was observed above 90°C, the enzyme regained more than 90% activity after boiling for 5 min and subsequent cooling at 37°C for 45 minutes. The enzyme was also evaluated for any cellulose activity and as expected, no detectable cellulase activity was observed.Figure 3

Bottom Line: The enzyme did not show any cellulose activity and was active under wide range of temperature (40°C to 80°C) and pH (4 to 9).The enzyme showed considerable thermo stability and regained over 90% of activity, when returned to 55°C after boiling for 5 min.These physiochemical properties of B. brevis xylanse show high potential of its applications in paper and pulp industry.

View Article: PubMed Central - PubMed

Affiliation: Amity Institute of Biotechnology, Amity University Rajasthan, Jaipur, 302001 India.

ABSTRACT
Xylanase gene isolated from Bacillus brevis was expressed in E. coli BL21. Sequencing of the gene (Gen Bank accession number: HQ179986) showed that it belongs to family 11 xylanases. The recombinant xylanase was predominantly secreted to culture medium and showed mesophilic nature (optimum activity at 55°C and pH 7.0). The cell free culture medium exhibited 30 IU/ml xylanse activity. The enzyme did not show any cellulose activity and was active under wide range of temperature (40°C to 80°C) and pH (4 to 9). The enzyme showed considerable thermo stability and regained over 90% of activity, when returned to 55°C after boiling for 5 min. These physiochemical properties of B. brevis xylanse show high potential of its applications in paper and pulp industry.

No MeSH data available.