ALBERT

Description: A novel esterase gene selected from metagenomic sequences of deep-sea hydrothermal vents was successfully expressed in Escherichia coli . The recombinant protein (est-OKK), which belongs to the lipolytic enzyme family V, exhibited high activity toward pNP-esters with short acyl chains and especially p-nitrophenyl butyrate. Site-mutagenesis results confirmed that est-OKK contains the nonclassical catalytic tetrad predicted by alignment and computational modeling. The est-OKK protein is a moderately thermophilic enzyme that is relatively thermostable, and highly salt-tolerant, which remained stable in 3 mol/L NaCl for 6 hr. The est-OKK protein showed the considerable alkalistability, displayed optimal activity at pH 9.0 and maintained approximately 70% of its residual activity after incubation at pH 10 for 4 hr. Furthermore, the est-OKK activity was strongly resistant to a variety of metal ions such as Co 2+ , Zn 2+ , Fe 2+ , Na + , and K + ; nonionic detergents such as Tween-20, Tween-80; and organic solvents such as acetone and isopropanol. Taken together, the novel esterase with unique characteristics may give us a new insight into the family V of lipolytic enzymes, and could be a highly valuable candidate for biotechnological applications such as organic synthesis reactions or food and pharmaceutical industries. We isolated a novel esterase from deep-sea hydrothermal vents using a metagenomic sequence-based strategy. This new enzyme is a moderately thermophilic enzyme that is relatively thermostable and highly salt-tolerant, and exhibits remarkable alkalistability and very good stability against a variety of metal ions, nonionic detergents and organic solvents.