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

• Published in: Trees (Berlin, West) West), 2020-06, Vol.34 (3), p.835-843
• Main Authors: Alper, Meltem, Öztetik, Elif, Kaya, M. Yaşar, Köçkar, Feray
• Format: Article
• Language: English
• Subjects: Agriculture; Biomedical and Life Sciences; Forestry; Functional Genomics; Life Sciences; Original Article; Plant Anatomy/Development; Plant Pathology; Plant Physiology; Plant Sciences
• ISSN: 0931-1890; 1432-2285
• DOI: 10.1007/s00468-020-01962-3

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 Pb GST Tau enzyme were determined. 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 ( Pb GST Tau) from Pinus brutia Ten. The full length Pb GST Tau ORF was 687 bp having a molecular mass of 27.37 kDa. Catalytic and ligand binding sites of Pb GST 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 K m of 3.66 mM and 0.3 mM, respectively. Pb GST 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 Pb GST Tau gene and the enzyme. This study is going to provide new insights into the phylogenetic and biochemical analysis of GST family in conifers.