Characterization of pullulanase type II from Bacillus cereus H1.5
Problem statement: Pullulanase is one of the important enzymes in starch industry. Search for the pullulanase with distinct features, possibly from easily grown bacterium, is of interest for industrial applications Approach: The extracellular pullulanase produced by Bacillus cereus HI.5 was purified...
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| Format: | Article |
| Language: | English |
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Science Publications
2009
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| Online Access: | http://psasir.upm.edu.my/id/eprint/13550/ http://psasir.upm.edu.my/id/eprint/13550/1/ajbbsp.2009.170.179.pdf |
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| author | Hii, Siew Ling Ling, Tau Chuan Mohamad, Rosfarizan Ariff, Arbakariya |
| author_facet | Hii, Siew Ling Ling, Tau Chuan Mohamad, Rosfarizan Ariff, Arbakariya |
| author_sort | Hii, Siew Ling |
| building | UPM Institutional Repository |
| collection | Online Access |
| description | Problem statement: Pullulanase is one of the important enzymes in starch industry. Search for the pullulanase with distinct features, possibly from easily grown bacterium, is of interest for industrial applications Approach: The extracellular pullulanase produced by Bacillus cereus HI.5 was purified by chromatographic method of DEAE-Sepharos
e, followed by Superdex gel filtration. The enzyme was characterized in terms of the optimal pH and temperature for activity as well as substrate specificity. Results:
The enzyme showed optimal activity at 55°C and pH 6.0. The thermostability and the thermoactivity of the enzyme were increased considerably in the presence of Ca2+. In the present of 2 mM Ca2+, the enzyme had half-life duration of more than 2 h at 50°C. Almost all metal ions had a strong inhibitory effect, except Ca2+ and Mn2+. The Ca2+ had a very strong stimulating effect on the enzyme, increasing its activity by 170%. The enzyme was activated by 2-mercaptoethanol and dithiothreitol, where as N-bromosuccinimide and Schardinger dextrins were inhibitors, suggesting that tryptophan and thiol residues may be important for the activity. The apparent Km and Vmax
value for pullulan was 1.1 mg mL−1 and 0.275 μmol min−1, respectively. A relative substrate specificity for
hydrolysis of pullulan, amylopectin and soluble starch by this pullulanase was 100%, 28.5% and 20.4%, respectively.
Conclusion: The enzyme was able to attack specifically the
α-1,6 linkages in pullulan to generate maltotriose as the major end product, as well as the α-1,4 linkages in amylopectin and soluble starch leading to the formation of a mixture of maltose and glucose and therefore be classified as a type II pullulanase or an amylopullulanase. |
| first_indexed | 2025-11-15T07:54:26Z |
| format | Article |
| id | upm-13550 |
| institution | Universiti Putra Malaysia |
| institution_category | Local University |
| language | English |
| last_indexed | 2025-11-15T07:54:26Z |
| publishDate | 2009 |
| publisher | Science Publications |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | upm-135502017-12-04T04:25:48Z http://psasir.upm.edu.my/id/eprint/13550/ Characterization of pullulanase type II from Bacillus cereus H1.5 Hii, Siew Ling Ling, Tau Chuan Mohamad, Rosfarizan Ariff, Arbakariya Problem statement: Pullulanase is one of the important enzymes in starch industry. Search for the pullulanase with distinct features, possibly from easily grown bacterium, is of interest for industrial applications Approach: The extracellular pullulanase produced by Bacillus cereus HI.5 was purified by chromatographic method of DEAE-Sepharos e, followed by Superdex gel filtration. The enzyme was characterized in terms of the optimal pH and temperature for activity as well as substrate specificity. Results: The enzyme showed optimal activity at 55°C and pH 6.0. The thermostability and the thermoactivity of the enzyme were increased considerably in the presence of Ca2+. In the present of 2 mM Ca2+, the enzyme had half-life duration of more than 2 h at 50°C. Almost all metal ions had a strong inhibitory effect, except Ca2+ and Mn2+. The Ca2+ had a very strong stimulating effect on the enzyme, increasing its activity by 170%. The enzyme was activated by 2-mercaptoethanol and dithiothreitol, where as N-bromosuccinimide and Schardinger dextrins were inhibitors, suggesting that tryptophan and thiol residues may be important for the activity. The apparent Km and Vmax value for pullulan was 1.1 mg mL−1 and 0.275 μmol min−1, respectively. A relative substrate specificity for hydrolysis of pullulan, amylopectin and soluble starch by this pullulanase was 100%, 28.5% and 20.4%, respectively. Conclusion: The enzyme was able to attack specifically the α-1,6 linkages in pullulan to generate maltotriose as the major end product, as well as the α-1,4 linkages in amylopectin and soluble starch leading to the formation of a mixture of maltose and glucose and therefore be classified as a type II pullulanase or an amylopullulanase. Science Publications 2009 Article PeerReviewed application/pdf en http://psasir.upm.edu.my/id/eprint/13550/1/ajbbsp.2009.170.179.pdf Hii, Siew Ling and Ling, Tau Chuan and Mohamad, Rosfarizan and Ariff, Arbakariya (2009) Characterization of pullulanase type II from Bacillus cereus H1.5. American Journal of Biochemistry and Biotechnology, 5 (4). pp. 170-179. ISSN 1553-3468; ESSN: 1558-6332 http://thescipub.com/html/10.3844/ajbbsp.2009.170.179 10.3844/ajbbsp.2009.170.179 |
| spellingShingle | Hii, Siew Ling Ling, Tau Chuan Mohamad, Rosfarizan Ariff, Arbakariya Characterization of pullulanase type II from Bacillus cereus H1.5 |
| title | Characterization of pullulanase type II from Bacillus cereus H1.5 |
| title_full | Characterization of pullulanase type II from Bacillus cereus H1.5 |
| title_fullStr | Characterization of pullulanase type II from Bacillus cereus H1.5 |
| title_full_unstemmed | Characterization of pullulanase type II from Bacillus cereus H1.5 |
| title_short | Characterization of pullulanase type II from Bacillus cereus H1.5 |
| title_sort | characterization of pullulanase type ii from bacillus cereus h1.5 |
| url | http://psasir.upm.edu.my/id/eprint/13550/ http://psasir.upm.edu.my/id/eprint/13550/ http://psasir.upm.edu.my/id/eprint/13550/ http://psasir.upm.edu.my/id/eprint/13550/1/ajbbsp.2009.170.179.pdf |