ScienceAsia 40(2014): 405-413 |doi:
Cloning, characterization, and heterologous expression of a dextranase gene from Penicillium˙pinophilum SMCU3-14
Panan˙Rerngsamran*, Pimonrut˙Temjitpukdee, Nilnate˙Assavasirijinda, Supat˙Chareonpornwattana, Suthep˙Thaniyavarn
ABSTRACT: 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 ∼66˙kDa), with a putative N-terminal 20-amino acid signal peptide, giving a predicted mature protein of 588 amino acids (MW of ∼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.
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|Department˙of˙Microbiology, Faculty˙of˙Science, Chulalongkorn˙University, Payathai, Bangkok˙10330˙Thailand
* Corresponding author, E-mail: firstname.lastname@example.org
Received 31 Mar 2014, Accepted 11 Nov 2014