TY - JOUR
T1 - Cloning, expression and characterization of theta-class glutathione S-transferase from the silkworm, Bombyx mori
AU - Yamamoto, Kohji
AU - Zhang, Pingbo
AU - Miake, Fumio
AU - Kashige, Nobuhiro
AU - Aso, Yoichi
AU - Banno, Yutaka
AU - Fujii, Hiroshi
N1 - Funding Information:
We are grateful to Dr. Toru Shimada at University of Tokyo and Dr. Kazuei Mita at National Institute of Agrobiological Sciences for supplying plasmid DNA. The authors are greatly indebted to Mr. Kei-ichi Makizumi at The Chemo-Sero-Therapeutic Research Institute (Kaketsuken), Kumamoto, Japan for amino acid sequence analysis. This work is supported by the National Bio-resources project (RR2002 for the silkworm) of the Ministry of Education, Science, and Culture of Japan.
PY - 2005/7
Y1 - 2005/7
N2 - This study focused on glutathione S-transferase (GST), one of the detoxification enzymes, from the silkworm, Bombyx mori (GSTT1). A cDNA encoding a putative GST was amplified by reverse transcriptase-polymerase chain reaction and sequenced. The deduced amino acid sequence revealed 59%, 57% and 56% identities to theta-class GSTs of Musca domestica, Anopheles gambiae and Drosophila melanogaster, respectively. GSTT1 was also estimated to be close to those GSTs in a phylogenetic tree. Recombinant GST (rGSTT1) was functionally overexpressed in Escherichia coli in a soluble form, purified to homogeneity, and characterized. The pH-optimum of rGSTT1 was broad from pH 4 to 9 and rGSTT1 retained more than 75% of its original activity after incubation at pH 5-11. Incubation for 30 min at temperatures below 50°C also affected the activity insignificantly. The Michaelis constant for 1-chloro-2,4-dinitrobenzene was 0.48 mM.
AB - This study focused on glutathione S-transferase (GST), one of the detoxification enzymes, from the silkworm, Bombyx mori (GSTT1). A cDNA encoding a putative GST was amplified by reverse transcriptase-polymerase chain reaction and sequenced. The deduced amino acid sequence revealed 59%, 57% and 56% identities to theta-class GSTs of Musca domestica, Anopheles gambiae and Drosophila melanogaster, respectively. GSTT1 was also estimated to be close to those GSTs in a phylogenetic tree. Recombinant GST (rGSTT1) was functionally overexpressed in Escherichia coli in a soluble form, purified to homogeneity, and characterized. The pH-optimum of rGSTT1 was broad from pH 4 to 9 and rGSTT1 retained more than 75% of its original activity after incubation at pH 5-11. Incubation for 30 min at temperatures below 50°C also affected the activity insignificantly. The Michaelis constant for 1-chloro-2,4-dinitrobenzene was 0.48 mM.
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U2 - 10.1016/j.cbpc.2005.04.012
DO - 10.1016/j.cbpc.2005.04.012
M3 - Article
C2 - 15950511
AN - SCOPUS:20544431934
VL - 141
SP - 340
EP - 346
JO - Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology
JF - Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology
SN - 0305-0491
IS - 3
ER -