Original Paper

World Journal of Microbiology and Biotechnology

, Volume 27, Issue 4, pp 841-850

First online:

Molecular cloning and expression of an extracellular α-amylase gene from an Antarctic deep sea psychrotolerant Pseudomonas stutzeri strain 7193

  • Jinwei ZhangAffiliated withKey Laboratory of Marine Biogenetic Resources, State Oceanic AdministrationThird Institute of Oceanography, State Oceanic AdministrationDove Marine Laboratory, School of Marine Science and Technology, Newcastle University Upon Tyne
  • , Runying ZengAffiliated withKey Laboratory of Marine Biogenetic Resources, State Oceanic AdministrationThird Institute of Oceanography, State Oceanic Administration Email author 

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Abstract

Psychrotolerant Pseudomonas stutzeri strain 7193 capable of producing an extracellular α-amylase was isolated from deep sea sediments of Prydz Bay, Antarctic. The 59678-Da protein (AmyP) was encoded by 1665-bp gene (amyP). The deduced amino acid sequence was identified with four regions, which are conserved in amylolytic enzymes and form a catalytic domain, and was predicted to be maltotetraose forming extracellular amylase by using the I-TASSER online server. Purification of AmyP amylases from both the recombinant of Escherichia coli Top 10 F′ and strain 7193 was conducted. Biochemical characterization revealed that the optimal amylase activity was observed at pH 9.0 and temperature 40°C. The enzymes were unstable at temperatures above 30°C, and only retain half of their highest activity after incubation at 60°C for 5 min. Thin-layer chromatography analysis of the products of the amylolytic reaction showed the presence of maltotetraose, maltotriose, maltose and glucose in the starch hydrolysate.

Keywords

Deep sea sediment Extracellular α-amylase Sequencing and expression Characterization