Abstract:
Microbulbifer strain CMC-5 was isolated from decomposing seaweeds, and was found to degrade agar, alginate, carboxymethyl cellulose, carrageenan, xylan, and chitin. The extracellular agarase enzyme from strain CMC-5 was purified 103-fold by ultrafiltration, ion-exchange chromatography, using diethylaminoethyl sepharose FF, and gel filtration, using sephacryl S-300HR, with a yield of 6.7 percent. Zymogram and protein staining of the purified agarase on a SDS-polyacrylamide gel revealed a single band, with an apparent molecular weight of 59 kDa. The purified enzyme was endo-type beta-agarase, as it was able to hydrolyze the beta-1,4 glycosidic linkages of agarose, releasing neoagarotetraose and neoagarohexaose as the end products. The optimum pH and temperature of agarase were 7 and 50 degrees C, respectively. Thermal stability studies indicated that the agarase retained 62 percent of its activity after incubating at 50 degrees C for 30 min. Treatment with EDTA reduced the agarase activity by 54 percent. The agarase activity was stimulated by the presence of Ca(2+) and Mg(2+) ions; whereas, Zn(2+), Hg(2+), Cu(2+), Fe(2+), and Co(2+) abolished the activity. Further, the presence of NaCl at concentrations lower than 100 mM caused a decrease in the agarase activity; whereas, the activity was enhanced up to a concentration of 500 mM.
