Expression, purification, and molecular analysis of the Necator americanus glutathione S-transferase 1 (Na-GST-1): a production process developed for a lead candidate recombinant hookworm vaccine antigen

Protein Expr Purif. 2012 Jun;83(2):145-51. doi: 10.1016/j.pep.2012.03.013. Epub 2012 Apr 4.


The enzyme Necator americanus glutathione S-transferase 1 (Na-GST-1) belongs to a unique Nu class of GSTs and is a lead candidate antigen in a bivalent human hookworm vaccine. Here we describe the expression of Na-GST-1 in the yeast Pichia pastoris at the 20 L manufacturing scale and its purification process performed by three chromatographic steps, comprised of a Q Sepharose XL anion exchange column, followed by a Butyl Sepharose HP hydrophobic affinity column and a Superdex 75 size-exclusion column. Approximately 1.5 g of recombinant protein was recovered at an overall process yield of 51%, with a purity grade of 98% and the absence of detectable host cell protein. By mass spectrometry the recombinant protein exhibits a mass of 23,676Da, which closely matches the predicted molecular mass of the protein. The expression and purification methods described here are suitable for further scale-up product development and for its use to design formulation processes suitable to generate a vaccine for clinical testing.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antigens, Helminth / genetics
  • Antigens, Helminth / isolation & purification*
  • Antigens, Helminth / metabolism
  • Chromatography, Gel
  • Chromatography, High Pressure Liquid
  • Electrophoresis, Polyacrylamide Gel
  • Glutathione Transferase / genetics
  • Glutathione Transferase / isolation & purification*
  • Glutathione Transferase / metabolism
  • Helminth Proteins / genetics
  • Helminth Proteins / isolation & purification*
  • Helminth Proteins / metabolism
  • Necator americanus / enzymology*
  • Pichia / genetics
  • Recombinant Proteins / genetics
  • Recombinant Proteins / isolation & purification*
  • Recombinant Proteins / metabolism


  • Antigens, Helminth
  • Helminth Proteins
  • Recombinant Proteins
  • Glutathione Transferase