The eukaryotic elongation factor 1 gamma (eEF1 gamma) is a multi-domain protein, which consist of a glutathione transferase (GST)-like N-terminus domain. In association with alpha, beta and delta subunits, eEF1 gamma forms part of the eukaryotic elongation factor complex, which is mainly involved in protein biosynthesis. The N-terminus GST domain of eEF1 gamma interacts with the beta subunit. eEF1 gamma subunit is over-expressed in human carcinoma. The role of human eEF1 gamma (heEF1 gamma) is poorly understood. A successful purification of recombinant heEF1 gamma is the first step towards determining unknown properties of the protein, including putative GST-like activities and the structure of the protein. This paper describes the over-expression, purification and characterisation of recombinant full-length, and the N- and C-terminus domains of heEF1 gamma. All three recombinant heEF1 gamma constructs over-expressed in the soluble Escherichia coil cell fraction and were purified to homogeneity. Secondary structure analysis indicates that the heEF1 gamma constructs have high alpha-helical structural character. The full-length and N-terminus domain are dimeric, while the C-terminus is monomeric. Both full-length and N-terminus domain interact with 8-anilino-1-naphthalene sulfonate (ANS) with K-D = 70.0 (+/- 5.7) mu M and with reduced glutathione (GSH). Glutathione sulfonate displaced ANS bound to hydrophobic binding sites in the recombinant N-terminus domain. Using the standard GSH-1-chloro-2,4-dinitrobenzene conjugation assay, the N-domain showed some enzyme activity (0.03 mu mol min(-1) mg(-1) protein), while the full-length heEF1 gamma did not catalyse the GSH-CDNB conjugation. Consequently, we hypothesize the presence of a presumed GST-like active site structure in the heEF1 gamma, which comprises a glutathione binding site and a hydrophobic substrate binding site. (C) 2014 Elsevier Inc. All rights reserved.