Purification and Characterization of Thermostable Agarase from Bacillus sp BI-3, a Thermophilic Bacterium Isolated from Hot Spring

An extracellular agarase was purified from Bacillus sp. BI-3, a thermophilic agar-degrading bacterium isolated from a hot spring in Indonesia. The purified agarase revealed a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, with an apparent molecular mass of 58 kDa. The optimum pH and temperature of the agarase were 6.4 and 70 degrees C, respectively. The activity of the agarase was stable at high temperatures, and more than 50% activity was retained at 80 degrees C for 15 min. Furthermore, the enzyme was stable in the pH range of 5.8-8.0, and more than 60% of the residual activity was retained. Significant activation of the agarase was observed in the presence of K+, Na+, Ca2+, Mg2+, and Sr2+; on the other hand, Ba2+, Zn2+, Cu2+, Mn2+, Co2+, Fe2+, and EDTA inhibited or inactivated the enzyme activity. The components of the hydrolytic product analyzed by thin-layer chromatography showed that the agarase mainly produced neoagarobiose. This study is the first to present evidence of agarolytic activity in aerobic thermophilic bacteria.