Journal of Industrial Microbiology & Biotechnology

, Volume 40, Issue 5, pp 437–446

Purification, characterization and cloning of a thermotolerant isoamylase produced from Bacillus sp. CICIM 304

  • Youran Li
  • Dandan Niu
  • Liang Zhang
  • Zhengxiang Wang
  • Guiyang Shi
Biocatalysis

DOI: 10.1007/s10295-013-1249-7

Cite this article as:
Li, Y., Niu, D., Zhang, L. et al. J Ind Microbiol Biotechnol (2013) 40: 437. doi:10.1007/s10295-013-1249-7

Abstract

A novel thermostable isoamylase, IAM, was purified to homogeneity from the newly isolated thermophilic bacterium Bacillus sp. CICIM 304. The purified monomeric protein with an estimated molecular mass of 100 kDa displayed its optimal temperature and pH at 70 °C and 6.0, respectively, with excellent thermostability between 30 and 70 °C and pH values from 5.5 to 9.0. Under the conditions of temperature 50 °C and pH 6.0, the Km and Vmax on glycogen were 0.403 ± 0.018 mg/mg and 0.018 ± 0.001 mg/(min mg), respectively. Gene encoding IAM, BsIam was identified from genomic DNA sequence with inverse PCRs. The open reading frame of the BsIam gene was 2,655 base pairs long and encoded a polypeptide of 885 amino acids with a calculated molecular mass of 101,155 Da. The deduced amino acid sequence of IAM shared less than 40 % homology with that of microbial isoamylase ever reported, which indicated it was a novel isoamylase. This enzyme showed its obvious superiority in the industrial starch conversion process.

Keywords

Bacillus sp.Thermostable isoamylasePurificationEnzyme propertiesCloning

Supplementary material

10295_2013_1249_MOESM1_ESM.doc (329 kb)
Supplementary material 1 (DOC 329 kb)

Copyright information

© Society for Industrial Microbiology and Biotechnology 2013

Authors and Affiliations

  • Youran Li
    • 1
  • Dandan Niu
    • 1
  • Liang Zhang
    • 1
  • Zhengxiang Wang
    • 1
  • Guiyang Shi
    • 1
  1. 1.Research Center of Bioresource & Bioenergy, School of BiotechnologyJiangnan UniversityWuxiPeople’s Republic of China