A Novel GH-50 Family β-agarase from Saccharophagus sp. AG21: Molecular Cloning and Enzymatic Characterization

Abstract

The agarase producing marine bacteria was isolated from seawater of Jeju Island. 16s rRNA sequence showed 99% (1376/1384 bp) identity with Saccharophagus degradans 2-40. The ORF sequence was amplified and cloned into pMAL-c2x expression vector. Then recombinant agarase was overexpressed and purified using E. coli expression system. The Hydrolized product of recombinant agarase was determined by thin layer chromatography. The 2334 base pair ORF of beta-agarase is encoding 778 amino acid residues. The predicted molecular mass was 88 kDa and isoelectric porint was 5.2. Pairwise amino acid comparison of agarase showed 96% identity with Saccharophagus degradans 2-40 agarase. The purified recombinant agarase was not able to degrade agar, however it could only degrade the neoagarotetraose and neoagarohexaose, mainly into neoagarobiose components. Supported by grant from KIOST and MLTM (PE98931, PM57210, PE98932). vector. Then recombinant agarase was overexpressed and purified using E. coli expression system. The Hydrolized product of recombinant agarase was determined by thin layer chromatography. The 2334 base pair ORF of beta-agarase is encoding 778 amino acid residues. The predicted molecular mass was 88 kDa and isoelectric porint was 5.2. Pairwise amino acid comparison of agarase showed 96% identity with Saccharophagus degradans 2-40 agarase. The purified recombinant agarase was not able to degrade agar, however it could only degrade the neoagarotetraose and neoagarohexaose, mainly into neoagarobiose components. Supported by grant from KIOST and MLTM (PE98931, PM57210, PE98932).