title: [Structure and function of a novel thermostable pullulanase].
abstract: Research on novel pullulanase has major significance on the domestic industrialization of pullulanase and the breakdown of foreign monopoly. A thermophilic bacteria LM 18-11 producing thermostable pullulanase was isolated from Lunma hot springs of Yunnan province. It was identified as Anoxybacillus sp. by 16S rDNA phylogenetic analysis. Full-length pullulanase gene was cloned from Anoxybacillus sp. LM18-11. The optimum temperature of the pullulanase was between 55 and 60 degrees C with a half-life as long as 48 h at 60 degrees C; and its optimum pH was between 5.6 and 6.4. V(max) and K(m) of the pullulanase was measured as 750 U/mg and 1.47 mg/mL, which is the highest specific activity reported so far. The pullulanase crystals structure showed a typical alpha-amylase family structure. The N-terminal has a special substrate binding domain. Activity and substrate binding were decreased when the domain was deleted, the V(max) and K(m) were 324 U/mg and 1.95 mg/mL, respectively. The pullulanase was highly heterologous expressed in Bacillus subtilis by P43 promoter. The extracellular enzyme activity was 42 U/mL, which increased more than 40 times compared to the initial strain. This pullulanase has good application prospects.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhen
authors.forename: Zheng
authors.initials: Z
authors.lastname: Hu
authors.forename: Shufang
authors.initials: S
authors.lastname: Li
authors.forename: Jianyong
authors.initials: J
authors.lastname: Xu
authors.forename: Hui
authors.initials: H
authors.lastname: Song
mesh_terms: D058677:Anoxybacillus / Q000145:classification / Q000201:enzymology*; D002681:China; D006026:Glycoside Hydrolases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D010802:Phylogeny; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D013696:Temperature
chemical_list: D012336:RNA, Ribosomal, 16S; D006026:Glycoside Hydrolases; C100181:pullulanase