Journal
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
Volume 60, Issue 44, Pages 11164-11172Publisher
AMER CHEMICAL SOC
DOI: 10.1021/jf303109u
Keywords
Bacillus sp.; thermostable type I pullulanase; gene cloning and expression
Funding
- China-South Africa Joint project [2009DFA31300]
- '863' program [2011AA100905]
- Program for New Century Excellent Talents in University [NCET-11-0665]
- Priority Academic Program Development of Jiangsu Higher Education Institutions
- 111 Project [111-2-06]
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The pulA1 gene, encoding a novel thermostable type I pullulanase PulA1 from Bacillus sp. CICIM 263, was identified from genomic DNA. The open reading frame of the pulA1 gene was 2655 base pairs long and encoded a polypeptide (PulA1) of 885 amino acids with a calculated molecular mass of 100,887 Da. The pulA1 gene was expressed in Escherichia coli and Bacillus subtilis. Recombinant PuLA1 showed optimal activity at pH 6.5 and 70 degrees C. The enzyme demonstrated moderate thermostability as PuLA1 maintained more than 88% of its acitivity when incubated at 70 degrees C for 1 h. The enzyme could completely hydrolyze pullulan to maltotriose, and hydrolytic activity was also detected with glycogen, starch and amylopection, but not with amylose, which is consistent with the property of type I pullulanase. PulA1 may be suitable for industrial applications to improve the yields of fermentable sugars for bioethanol production.
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