Cloning, expression, purification and preliminary X-ray analysis of EstN2, a novel archaeal α/β-hydrolase from Candidatus Nitrososphaera gargensis

Acta Crystallogr F Struct Biol Commun. 2014 Oct;70(Pt 10):1394-7. doi: 10.1107/S2053230X14018482. Epub 2014 Sep 25.

Abstract

EstN2 is a novel α/β-hydrolase originating from the ammonia-oxidizing thaumarchaeon Candidatus Nitrososphaera gargensis. The genome of the organism was sequenced and genes conferring putative lipolytic activity were amplified and cloned into Escherichia coli as a heterologous host. Through function-based screening, esterase and lipase activity was detected. A recombinant enzyme designated EstN2 was successfully expressed, purified and crystallized. The crystals belonged to space group I2, with one molecule per asymmetric unit, and diffracted X-rays to 1.5 Å resolution.

Keywords: Candidatus Nitrososphaera gargensis; EstN2; α/β-hydrolase.

MeSH terms

  • Amino Acid Sequence
  • Archaea / enzymology*
  • Archaeal Proteins / biosynthesis
  • Archaeal Proteins / chemistry*
  • Archaeal Proteins / isolation & purification
  • Chromatography, Affinity
  • Cloning, Molecular
  • Crystallization
  • Crystallography, X-Ray
  • Escherichia coli
  • Gene Expression
  • Hydrolases / biosynthesis
  • Hydrolases / chemistry*
  • Hydrolases / isolation & purification
  • Molecular Sequence Data

Substances

  • Archaeal Proteins
  • Hydrolases