Extremophiles

, Volume 9, Issue 4, pp 297–305

A thermostable phosphotriesterase from the archaeon Sulfolobus solfataricus: cloning, overexpression and properties

  • Luigia Merone
  • Luigi Mandrich
  • Mosè Rossi
  • Giuseppe Manco
Original Paper

DOI: 10.1007/s00792-005-0445-4

Cite this article as:
Merone, L., Mandrich, L., Rossi, M. et al. Extremophiles (2005) 9: 297. doi:10.1007/s00792-005-0445-4

Abstract

A new gene from the hyperthermophilic archaeon Sulfolobus solfataricus MT4, coding for a putative protein reported to show sequence identity with the phosphotriesterase-related protein family (PHP), was cloned by means of the polymerase chain reaction from the S. solfataricus genomic DNA. In order to analyse the biochemical properties of the protein an overexpression system in Escherichia coli was established. The recombinant protein, expressed in soluble form at 5 mg/l of E. coli culture, was purified to homogeneity and characterized. In contrast with its mesophilic E. coli counterpart that was devoid of any tested activity, the S. solfataricus enzyme was demonstrated to have a low paraoxonase activity. This activity was dependent from metal cations with Co2+, Mg2+ and Ni2+ being the most effective and was thermophilic and thermostable. The enzyme was inactivated with EDTA and o-phenantroline. A reported inhibitor for Pseudomonas putida phosphotriesterase (PTE) had no effect on the S. solfataricus paraoxonase. The importance of a stable paraoxonase for detoxification of chemical warfare agents and agricultural pesticides will be discussed.

Keywords

PhosphotriesteraseOrganophosphatesPesticidesSulfolobus solfataricusThermostability

Abbreviations

PTE

Phosphotriesterase

OPH

Organophosphate hydrolase

OPD

Organo-phosphate-degrading enzyme

OPs

Organophosphates

SsoPox

Sulfolobus solfataricus paraoxonase

ePHP

E. coli phosphotriesterase-related protein

pNP

P-nitrophenyl

LB

Luria-Bertani

IPTG

Isopropyl-β-D-thiogalactopyranoside

PVDF

Polyvinylidene fluoride

EST2

Esterase 2

Copyright information

© Springer-Verlag 2005

Authors and Affiliations

  • Luigia Merone
    • 1
  • Luigi Mandrich
    • 1
  • Mosè Rossi
    • 1
  • Giuseppe Manco
    • 1
  1. 1.Istituto di Biochimica delle ProteineConsiglio Nazionale delle RicercheNapoliItaly