Molecular cloning, expression and characterization of glutathione S-transferase from Mytilus edulis.

The gene coding for glutathione S-transferase (GST) has been isolated from the Mytilus edulis hepatopancreas. Open reading frame analysis indicated that the M. edulis GST (meGST) gene encodes a protein of 206 amino acid residues with a calculated molecular mass of 23.68 kDa. The deduced amino acid sequence showed high sequence similarity with the sequence of the pi class GST. The meGST was expressed in Escherichia coli, and the recombinant meGST was purified by affinity chromatography and characterized. The recombinant meGST exhibited high activity towards the substrates ethacrynic acid (ECA) and 1-chloro-2,4-dinitrobenzene (CDNB). Kinetic analysis with respect to CDNB as substrate gave a K(m) of 0.68 mM and a V(max) of 0.10 mmol/min per mg protein. The recombinant meGST had a maximum activity at approximately pH 8.5, and its optimum temperature was 39 degrees C. The predicted three-dimensional structure of the meGST revealed the N-terminal domain possesses a thioredoxin fold and the six helices of the C-terminal domain make a alpha-helical bundle. These features indicate that the meGST belongs to pi class GST.