Molecular cloning and biochemical characterization of a Tau class glutathione S-transferase from Pinus brutia Ten

Alper, Meltem ( Aksaray, Yazar )Key message: A new Tau class GST gene was cloned from Pinus brutia Ten. cDNA sequence was analysed for conserved sequences. Substrate specificity, optimum pH, and temperature values of the recombinant PbGST Tau enzyme were determined. Abstract: Tau class glutathione S-transferases (GSTs) are essential enzymes for detoxification in plants. To date, a lot of the members of this family have been characterized from different plants but the studies on the conifers are very scarce. This study investigates for the first time molecular cloning and biochemical characterization of a Tau class GST gene (PbGST Tau) from Pinus brutia Ten. The full length PbGST Tau ORF was 687 bp having a molecular mass of 27.37 kDa. Catalytic and ligand binding sites of PbGST Tau are well conserved and shared maximum identity with Pinus tabulaeformis GST Tau. Kinetic analysis with respect to 1-chloro-2,4-dinitrobenzene (CDNB) and ethacrynic acid (ECA) as substrates exhibited a Km of 3.66 mM and 0.3 mM, respectively. PbGST Tau enzyme had an optimum activity at pH 6.0 and 8.0 when CDNB and ECA were used as substrate, respectively. The highest activity was measured at 25 °C. Through enzyme assays, phylogenetic analysis and structural modelling, we provide a detailed characterization of the PbGST Tau gene and the enzyme. This study is going to provide new insights into the phylogenetic and biochemical analysis of GST family in conifers

Novel debittering process of green table olives: application of β-glucosidase bound onto superparamagnetic nanoparticles

In this work, the olive β-glucosidase (β-glu) enzyme was immobilized onto superparamagnetic nanoparticles (SPMNs). Moreover, immobilized enzyme was also used for the debittering process for natural edible olives from Edremit, Turkey. Owing to SPMNs, the system can be easily removed by a simple magnet and reused many times for debittering process. The free olive β-glucosidase (E), β-glucosidase bound SPMNs (IE), free commercial β-glucosidase (CE), and commercial β-glucosidase bound SPMNs (ICE) were comparatively studied for oleuropein removal. In 6 h, the treatment of E, IE, and ICE hydrolyzed the 15.8%, 56.4%, and 80.0% of the oleuropein for 1000 g of olives, respectively, and further treatment showed that the IE and ICE reached the 100% after 22 h treatment. The results revealed that the IE and ICE with biocompatible properties of SPMNs have the economical, fast, and reusable properties for the industrial debittering process in the table olive production