Molecular cloning, expression, and characterization of a beta-agarase gene, agaD, from a marine bacterium, Vibrio sp. strain PO-303.
The beta-agarase-d gene (agaD) from a marine bacterium, Vibrio sp. strain PO-303, was cloned and expressed in Escherichia coli. The gene consists of 1,362 bp and encodes a protein of 453 amino acids with a predicted molecular weight of 50,824. The full length of agarase-d consists of a signal peptide, a glycoside hydrolase family 16 catalytic module (CM), and a carbohydrate binding module (CBM). The full length of agarase-d without the signal peptide (rAgaDDeltafull), the catalytic module (rAgaDCM), or the CBM (rAgaDCBM) was expressed in E. coli as recombinant proteins. rAgaDCM exhibited higher enzyme activity (63.6 units/mg) than rAgaDDeltafull (1.20 units/mg) against agarose. rAgaDCM hydrolyzed agar and porphyran to several oligosaccharides and acted on neoagarohexaose to produce neoagarotetraose and neoagarobiose, but did not act on neoagarotetraose. rAgaDCBM bound to agarose.
Dong
Jinhua
J
Graduate School of Bioresources, Mie University, Japan.
Tamaru
Yutaka
Y
Araki
Toshiyoshi
T
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017124:Conserved Sequence; D003470:Culture Media; D015870:Gene Expression; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D006868:Hydrolysis; D008969:Molecular Sequence Data; D009792:Oceans and Seas; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014733:Vibrio / Q000201:enzymology* / Q000235:genetics
D003470:Culture Media; D006026:Glycoside Hydrolases; C019624:agarase