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Gene Cloning and Characterization of Pcal_0222, α-Amylase from Pyrobaculum calidifontis

Gene Cloning and Characterization of Pcal_0222, α-Amylase from Pyrobaculum calidifontis

Sadaf Ashraf1, Masood Ahmed Siddiqui1*, Kanwal Nisa1, Samar Ali1 and Naeem Rashid2

1Department of Chemistry, Biotechnology Research Laboratory, University of Balochistan, Quetta 87300, Pakistan 
2School of Biological Sciences, University of the Punjab, Quaid-e-Azam Campus, Lahore 54590, Pakistan

*      Corresponding author: masoodbioeng@hotmail.com

ABSTRACT

The gene encoding Pcal_0222 from hyperthermophilic archaeon Pyrobaculum calidifontis was cloned and expressed in Escherichia coli. Pcal_0222 was composed of 529 amino acids with a theoretical molecular mass of 58 kDa. The amino acid sequence contained the four conserved regions that are a characteristic of GH13 family members. Recombinant Pcal_0222 was purified to apparent homogeneity using cation exchange and gel filtration column chromatographies. Purified Pcal_0222 exhibited optimal α-amylase activity at 85°C and pH 5.5. The activity was not dependent on any metal ion. The hyperthermophilic nature and metal ion independence make it a suitable candidate for both basic as well as applied research.
 

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Pakistan Journal of Zoology

October

Vol. 54, Iss. 5, Pages 2003-2500

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