title: Cloning, Expression, and Purification of Hyperthermophile α-Amylase from Pyrococcus woesei.
abstract: OBJECTIVE
  In an attempt α-amylase gene from Pyrococcus woesei was amplified and cloned into a pTYB2 vector to generate the recombinant plasmid pTY- α-amylase.
  
  
  METHODS
  Escherichia coli BL21 used as a host and protein expression was applied using IPTG. SDS-PAGE assay demonstrated the 100 kDa protein. Amylolytic activity of proteins produced by transformed E. coli cells was detected by zymography, and the rate of active α-amylase with and without the intein tag in both soluble conditions and as inclusion bodies solubilized by 4M urea were measured.
  
  
  RESULTS
  Amylolytic activity of ∼185,000 U/L of bacterial culture was observed from the soluble form of the protein using this system.
  
  
  CONCLUSIONS
  These results indicate that this expression system was appropriate for the production of thermostable α-amylase.
authors.affiliation: Department of Microbiology and Immunology, Faculty of Medicine, Kashan University of Medical Sciences, Kashan, Iran.
authors.forename: Amir
authors.initials: A
authors.lastname: Ghasemi
authors.affiliation: Clinical Microbiology Research Center, Ilam University of Medical Sciences, Ilam, Iran.
authors.forename: Sobhan
authors.initials: S
authors.lastname: Ghafourian
authors.affiliation: Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, Iran.
authors.forename: Sedighe
authors.initials: S
authors.lastname: Vafaei
authors.affiliation: Clinical Microbiology Research Center, Ilam University of Medical Sciences, Ilam, Iran.
authors.forename: Reza
authors.initials: R
authors.lastname: Mohebi
authors.affiliation: Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, Iran.
authors.forename: Maryam
authors.initials: M
authors.lastname: Farzi
authors.affiliation: Razi Herbal Medicines Research Center & Department of Microbiology, School of Medicine, Lorestan University of Medical Sciences, Khorramabad, Iran.
authors.forename: Morovat
authors.initials: M
authors.lastname: Taherikalani
authors.affiliation: Clinical Microbiology Research Center, Ilam University of Medical Sciences, Ilam, Iran.
authors.forename: Nourkhoda
authors.initials: N
authors.lastname: Sadeghifard
keywords: amylase; expression; recombinant protein; thermophile
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