Purification and characterization of glutathione S-transferase from needles of air polluted Scots pine (Pinus sylvestris L.) trees

The activity of glutathione S-transferase (GST, EC 2.5.1.18) was determined in needles from Scots pines (Pinus sylvestris L.) obtained from two polluted field sites. Substantial differences were found between the two sites' samples regarding the levels of soluble protein and the total GST activity in the crude extracts. GST was purified more than 90-fold using ammonium sulphate precipitation, gel filtration and affinity chromatography, resulting in 15 % recovery. The purified enzyme of both sites had a pH optimum between 7.75 and 8.0 and showed high specificity for, 1-chloro-2,4-dinitro-benzene (CDNB) as substrate while activities with 1,2-dichloro-4-nitrobenzene (DCNB) were much lower. The approximate molecular weight was determined as 54 kDa for the native enzyme and 26 kDa for the subunits. The enzyme of the two sampling sites differed in their affinity for glutathione and CDNB.