Activity optimisation of extracellular agarases produced by agarolytic bacteria Flammeovirga yaeyamensisAM5.A, Aliagarivorans marinusAM17. E1 and Aliagarivorans taiwanensisA69.B2, isolated from coral reef ecosystems

Abstract

The present study was undertaken to standardise the conditions for activity optimisation of agarases produced by three distinct strains of agar degrading bacteria viz, Flammeovirga yaeyamensisAM5.A, Aliagarivorans marinusAM17.E1 and Aliagarivorans taiwanensisA69.B2 (GenBank Acc. Nos. MT473965; MT475710 and MT473967 respectively) isolated from coral reef ecosystems along the southern coast of India. Agarase from F. yaeyamensis exhibited highest activity at 40°C, while A. marinus and A. taiwanensis agarases had peak activity at 50°C. The optimal pH and incubation time for agarases from all the three strains were 7.0 and 45 min respectively. The partially purified enzyme-extracts from the three strains were further studied for their responses to the presence of various metal ions (Cu2+, K+ , Hg2+ , Mn+ , Na+ and Ca2+); a chelating agent, ethylenediamine tetraacetic acid (EDTA); a reducing agent, mercaptoethanol and a serine protease inhibitor, phenyl methyl sulfonyl fluoride (PMSF). Enzyme characterisation results clearly indicated the sensitivity of all the three agarases to Hg2+ and Na+ ions. Cu2+ ions were found inhibitory to the enzyme from A. taiwanensis and A. marinus. However, F. yaeyamensis derived agarases remained unaffected even at 5 mM concentrations of Cu2+. Presence of K+ ions evidently suppressed the agarases from F. yaeyamensis and A. marinus. EDTA, mercaptoethanol and PMSF were found to be inhibitors of the enzyme activity, while Mn+ and Ca2+ had additive effect. The results of the study indicated potential of the bacterial strains investigated in this study as prospective sources of agarases.

Keywords: Agar degrading bacteria, Chelating agent, Metal ions, pH, Reducing agent, Serine protease inhibitor, Temperature, Time of incubation