Molecular and biochemical characterisation of arginine kinases in Haemonchus contortus and Teladorsagia circumcincta

Exp Parasitol. 2013 Jul;134(3):362-7. doi: 10.1016/j.exppara.2013.03.011. Epub 2013 Apr 6.

Abstract

Full length cDNA encoding arginine kinases (AK) were cloned from Teladorsagia circumcincta (TcAK) and Haemonchus contortus (HcAK). The TcAK and HcAK cDNA (1080 bp) encoded 360 amino acid proteins. The predicted amino acid sequence showed 99% similarity with each other and 94% with a Caenorhabditis elegans AK. Soluble N-terminal His-tagged AK proteins were expressed in Escherichia coli strain BL21, purified and characterised. All binding sites were completely conserved in both proteins. The recombinant TcAK and HcAK had very similar kinetic properties: K(m) arginine was 0.35 mM, K(m) ATP was 0.8-0.9 mM and the pH optima were pH 7.5. Arginine analogues strongly inhibited recombinant enzyme activities (up to 80%), whilst other amino acids decreased activities by a maximum of 20%. TcAK and HcAK are potential vaccine candidates because of the strong antigenicity of invertebrate phosphagens and kinases and presence in metabolically active parts of the worm.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Arginine / metabolism
  • Arginine Kinase / antagonists & inhibitors
  • Arginine Kinase / chemistry
  • Arginine Kinase / genetics*
  • Binding Sites
  • Cloning, Molecular
  • DNA, Complementary / genetics
  • DNA, Helminth / genetics
  • Electrophoresis, Polyacrylamide Gel
  • Haemonchus / enzymology*
  • Haemonchus / genetics
  • Helminth Proteins / chemistry
  • Helminth Proteins / genetics
  • Hydrogen-Ion Concentration
  • Kinetics
  • Molecular Sequence Data
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Sequence Alignment
  • Trichostrongyloidea / enzymology*
  • Trichostrongyloidea / genetics

Substances

  • DNA, Complementary
  • DNA, Helminth
  • Helminth Proteins
  • Recombinant Proteins
  • Arginine
  • Arginine Kinase

Associated data

  • GENBANK/JX422017
  • GENBANK/JX422018