Identification of a sigma‐class glutathione‐S‐transferase from the silkworm,Bombyx mori

Abstract

A cDNA that encodes a glutathione-S-transferase belonging to sigma class (GSTS1) from the silkworm, Bombyx mori was cloned by reverse transcriptase-polymerase chain reaction and sequenced. The deduced amino acid sequence revealed 66%, 48% and 41% identity to sigma-class GSTs from Manduca sexta, Blattella germanica and Anopheles gambiae, respectively. The GSTS1 was also estimated to be close to those GSTs in a phylogenetic tree. GSTS1 mRNA was widely distributed in various tissues. The recombinant GST (rGSTS1) was functionally overexpressed in Escherichia coli in a soluble form, purified to homogeneity and characterized. The pH-optimum of rGSTS1 was around pH 8. The rGSTS1 retained more than 75% of its original activity after incubation at pH 4-9. Incubation for 30 min at temperatures below 40°C did not affect its activity. rGSTS1 was able to catalyse the reaction of glutathione with 1-chloro-2,4-dinitrobenzene, the universal substrate for GST, as well as with 4-hydroxynonenal, the product of lipid peroxidation.

Biochemical characterization of an unclassified glutathione <i>S</i>-transferase of <i>Plutella xylostella</i>

“…4A), slightly lower than those of bmGSTD, bmGSTE, theta-class GST of B. mori (bmGSTT), and sigma-class GST of B. mori (bmGSTS), as well as an H. cunea sigma-class GST. 4,6,9,17,18) pxGSTu1 was stable at temperatures below 40°C (Fig. 4B), similar to bmGSTS, whereas bmGSTT, bmGSTE, bmGSTD, omega-class GST of B. mori (bmGSTO), and zeta-class GST of B. mori (bmGSTZ) were stable at temperatures below 50°C.…”

“…These sizes were similar to those of isolated lepidopteran GSTs. [3][4][5][6][7][8][9] Finally, we obtained 1.2 mg of highly purified pxGSTu1 from 250 mL of Luria-Bertani growth medium. The specific activity of the final preparation toward CDNB was 0.041 µmol/min/mg (Table 1).…”

“…1.18] are enzymes that are widespread in both prokaryotic and eukaryotic cells that catalyze the glutathione conjugation reaction with reduced glutathione (GSH). 1,2) We have identified several GSTs (delta, epsilon, omega, sigma, theta, zeta, and unclassified) in the silkworm, Bombyx mori, a lepidopteran model insect, [3][4][5][6][7][8][9] while six GST classes (delta, epsilon, omega, sigma, theta, and zeta) have been identified in dipteran insects, such as Anopheles gambiae 10) and Drosophila melanogaster. 11,12) In addition to the silkworm, we also characterized a sigma-class GST in the fall webworm Hyphantria cunea, one of the most serious lepidopteran pests of broad-leaved trees, 6) and a delta-class GST in Nilaparvata lugens, which is a rice crop pest.…”

Identification, genomic organization, and oxidative stress response of a sigma class glutathione S-transferase gene (AccGSTS1) in the honey bee, Apis cerana cerana

“…Finally, 4.8 mg of highly purified AccGSTS1 from 250 mL of LB medium was obtained. In previous studies, recombinant Sigma class GSTs from the fall webworm (Yamamoto et al 2007), silkworm (Yamamoto et al 2006), and earthworm (LaCourse et al 2009) exhibited specific activities for CDNB ranging between 1.9 and 4.4 μmolmin −1 mg −1 . In the present study, the specific activity of the final product toward CDNB was 2.1 μmolmin −1 mg −1 , revealing that active AccGSTS1 was successfully expressed in E. coli cells.…”

“…Because GST genes play important roles in detoxification pathways and the oxidative stress response, studies on GST resistance have been widely reported in insects, including Drosophila melanogaster Low et al 2007;Sawicki et al 2003;Tang and Tu 1994;Tu and Akgul 2005), Anopheles gambiae (Chen et al 2003;Ding et al 2003Ding et al , 2005Ranson et al 2000Ranson et al , 2001, and Bombyx mori (Yamamoto et al 2006;Yu et al 2008), but information about other important species is rather limited. Honey bees, which serve as an excellent model for social behavior research, are a major plant pollinator and thus play essential roles in agriculture and ecological balance.…”