Cloning and nucleotide sequence of the alpha-amylase gene from alkalophilic Pseudomonas sp KFCC 10818

Abstract

A gene coding for a new amylolytic enzyme from Pseudomonas sp. KFCC 10818 was cloned, and its nucleotide sequence was determined. Starting from an ATG initiation codon, there was an open reading frame composed of 1,398 bases in the sequence. A deduced amino acid sequence contained four highly conserved regions of alpha-amylases. Cloned amylase was purified from Escherichia coli, NH2-terminal sequencing of the enzyme showed the presence of a signal peptide composed of 23 amino acids. Maltose and maltotriose were major end products from starch by the enzyme action. pH and temperature optima of the alpha-amylase were pH 8 and 45 degrees C, respectively. The enzyme kept almost all catalytic activity during 3 h incubation between pH 7-11.