An extracellular halophilic protease SptA from a halophilic archaeon Natrinema sp. J7: Gene cloning, expression and characterization

Wanliang Shi, Xiao Feng Tang, Yuping Huang, Fei Gan, Bing Tang, Ping Shen

Research output: Contribution to journalArticlepeer-review

Abstract

A gene encoding an extracellular protease, sptA, was cloned from the halophilic archaeon Natrinema sp. J7. It encoded a polypeptide of 565 amino acids containing a putative 49-amino acid signal peptide, a 103-amino acid propeptide, as well as a mature region and C-terminal extension, with a high proportion of acidic amino acid residues. The sptA gene was expressed in Haloferax volcanii WFD11, and the recombinant enzyme could be secreted into the medium as an active mature form. The N-terminal amino acid sequencing and MALDI-TOF mass spectrometry analysis of the purified SptA protease indicated that the 152-amino acid prepropeptide was cleaved and the C-terminal extension was not processed after secretion. The SptA protease was optimally active at 50°C in 2.5 M NaCl at pH 8.0. The NaCl removed enzyme retained 20% of its activity, and 60% of the activity could be restored by reintroducing 2.5 M NaCl into the NaCl removed enzyme. When the twin-arginine motif in the signal peptide of SptA protease was replaced with a twin-lysine motif, the enzyme was not exported from Hfx. volcanii WFD11, suggesting that the SptA protease was a Tat-dependent substrate.

Original languageEnglish (US)
Pages (from-to)599-606
Number of pages8
JournalExtremophiles
Volume10
Issue number6
DOIs
StatePublished - Dec 2006
Externally publishedYes

Keywords

  • Extracellular protease
  • Halophilic archaeon
  • Natrinema sp. J7

ASJC Scopus subject areas

  • Microbiology
  • Molecular Medicine

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