Cloning, characterization, and heterologous expression of a dextranase gene from Penicillium pinophilum SMCU3-14

A DNA fragment encoding dextranase was cloned from Penicillium pinophilum SMCU3-14 using the genome walking approach. Sequence analysis of the gene (SMCU-DEX) revealed a putative CAAT box at 165 (CCAAT), a putative TATA box at 93 (TATAA) in the 5 '-noncoding region, and a polyadenylation signal (AATAAG) in the 3 '-noncoding region. A cDNA sequence analysis revealed no evidence of introns. The deduced open reading frame is 1824 bp in length and encodes a predicted protein of 608 amino acids (molecular weight (MW) of similar to 66 kDa), with a putative N-terminal 20-amino acid signal peptide, giving a predicted mature protein of 588 amino acids (MW of, similar to 64 kDa) that belongs to glycosyl hydrolase family 49, as with other fungal dextranases. This is the first report of a dextranase gene sequence from P. pinophilum. The cDNA was cloned and expressed in Escherichia coli, and the transformants showed dextranase activity on a dextran-containing agar medium. Crude extracts from the transformants analysed by sodium dodecyl sulphate polyacrylamide gel electrophoresis containing blue dextran revealed a distinct specific band of dextranase activity at an MW of approximately 66 kDa. This recombinant dextranase is likely to have valuable and cost-effective applications in medicine and industry.