Purification and characterisation of recombinant human eukaryotic elongation factor 1 gamma.

摘要: Abstract The eukaryotic elongation factor 1 gamma (eEF1γ) is a multi-domain protein, which consist of glutathione transferase (GST)-like N-terminus domain. In association with α, β and δ subunits, eEF1γ forms part the complex, mainly involved in protein biosynthesis. GST domain interacts subunit. subunit over-expressed human carcinoma. role (heEF1γ) poorly understood. A successful purification recombinant heEF1γ first step towards determining unknown properties including putative GST-like activities structure protein. This paper describes over-expression, characterisation full-length, N- C-terminus domains heEF1γ. All three constructs soluble Escherichia coli cell fraction were purified to homogeneity. Secondary analysis indicates that have high α-helical structural character. full-length are dimeric, while monomeric. Both interact 8-anilino-1-naphthalene sulfonate (ANS) KD = 70.0 (±5.7) μM reduced (GSH). Glutathione displaced ANS bound hydrophobic binding sites Using standard GSH-1-chloro-2,4-dinitrobenzene conjugation assay, N-domain showed some enzyme activity (0.03 μmol min−1 mg−1 protein), did not catalyse GSH-CDNB conjugation. Consequently, we hypothesize presence presumed active site heEF1γ, comprises substrate site.