Purification and Characterization of Glutathione S-Transferases of Eoreuma loftini and Diatraea saccharalis (Lepidoptera: Pyralidae)

Abstract

Clutathione S-transferases (CST) from larvae of the sugarcane borer, Diatraea Saccharalis (F.) and the Mexican rice borer, Eareuma laftini (Dyar), were characterized. Apparently homogeneous CST isoenzymes from the larvae of both species were purified with CSH-affinity chromatography and isoelectric focusing. E. laftini had higher CST activity (31.12 units/g wet weight of tissue) compared with D. saccharalis (24.8 units/g wet weight of tissue). The CST isoenzyme of D. saccharalis with a pi value of 9.3 was a dimer of two identical subunits ( M , 25,000), whereas the CST isoenzyme with a pi value of 8.0 was a heterodimer with subunit M, values of 25,000 and 27,000. The CST isoenzymes of E. laftini with pI values of 9.7 and 7.7 were homo dimers of subunits with M, values of 25,000 and 26,000, respectively. However, the CST isoenzyme with a pI value of 5.3 was a heterodimer of subunits having M, values of 26,000 and 27,000. Peptide fingerprint analysis by SV-8 protease digestion revealed primary structural differences in these isoenzymes. The N-termini of all CST isoenzymes from both the species of larvae were blocked. Substrate specificities and kinetic properties of the purified CSTs from both the species have been described. The low Km and high V max values toward l-chloro-2,4-dinitrobenzene of CST enzymes isolated from both D. saccharalis and E. laftini indicate that these enzymes have a high affinity for electrophilic substrates compared with most human CSTs.