title: Expression and biochemical characterization of a novel type I pullulanase from Bacillus megaterium.
abstract: OBJECTIVE
  To identify novel pullulanases from microorganisms and to investigate their biochemical characterizations.
  
  
  RESULTS
  A novel pullulanase gene (BmPul) from Bacillus megaterium WW1210 was cloned and heterologously expressed in Escherichia coli. The gene has an ORF of 2814 bp encoding 937 amino acids. The recombinant pullulanase (BmPul) was purified to homogeneity and biochemically characterized. BmPul has an MW of approx. 112 kDa as indicated by SDS-PAGE. Optimum conditions were at 55 °C and pH 6.5. The enzyme was stable below 40 °C and from pH 6.5-8.5. The Km values of BmPul towards pullulan and amylopectin were 3.3 and 3.6 mg/ml, respectively. BmPul hydrolyzed pullulan to yield mainly maltotriose, indicating that it should be a type I pullulanase.
  
  
  CONCLUSIONS
  A novel type I pullulanase from Bacillus megaterium was identified, heterologously expressed and biochemically characterized. Its properties makes this enzyme as a good candidate for the food industry.
authors.affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science & Nutritional Engineering, China Agricultural University, Beijing, 100083, China.
authors.forename: Shaoqing
authors.initials: S
authors.lastname: Yang
authors.affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, Bioresource Utilization Laboratory, College of Engineering, China Agricultural University, Beijing, 100083, China.
authors.forename: Qiaojuan
authors.initials: Q
authors.lastname: Yan
authors.affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, Bioresource Utilization Laboratory, College of Engineering, China Agricultural University, Beijing, 100083, China.
authors.forename: Qingdan
authors.initials: Q
authors.lastname: Bao
authors.affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science & Nutritional Engineering, China Agricultural University, Beijing, 100083, China.
authors.forename: Jingjing
authors.initials: J
authors.lastname: Liu
authors.affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science & Nutritional Engineering, China Agricultural University, Beijing, 100083, China. zhqjiang@cau.edu.cn.
authors.forename: Zhengqiang
authors.initials: Z
authors.lastname: Jiang
mesh_terms: D000595:Amino Acid Sequence; D000687:Amylopectin / Q000378:metabolism; D001410:Bacillus megaterium / Q000201:enzymology*; D002855:Chromatography, Thin Layer; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005798:Genes, Bacterial; D005936:Glucans / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D011994:Recombinant Proteins / Q000302:isolation & purification; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D005936:Glucans; D011994:Recombinant Proteins; C009109:pullulan; D000687:Amylopectin; D006026:Glycoside Hydrolases; C100181:pullulanase
keywords: Amylopectin; Bacillus megaterium; Characterization; Gene cloning; Pullulan; Pullulanase