A new phosphotriesterase cloned from the archaeon Sulfolobus solfataricus MT4

Abstract

A new gene from the hyperthermophilic archaeon Sulfolobus solfataricus MT4, reported to show sequence identity with members of the amidohydrolase superfamily, was cloned by means of the polymerase chain reaction from the S. solfataricus genomic DNA. In order to analyse the biochemical properties of this protein an overexpression system in Escherichia coli was established. The recombinant protein was purified to homogeneity and characterized. The S. solfataricus enzyme was demonstrated to have a low paraoxonase activity and also an even lower but detectable esterase activity. The paraoxonase activity was dependent from metal cations present in the E. coli culture with Cd²⁺>Ni²⁺=Co²⁺>Zn²⁺ and was thermophilic and thermostable. A 3D model of SsoPox was constructed on the basis of the Pseudomonas putida phosphotriesterase (PTE) structure and residues likely to be involved in substrate specificity and catalysis were identified.