TY - JOUR
T1 - Structural basis for catalytic activity of a silkworm Delta-class glutathione transferase
AU - Yamamoto, Kohji
AU - Usuda, Kazuhiro
AU - Kakuta, Yoshimitsu
AU - Kimura, Makoto
AU - Higashiura, Akifumi
AU - Nakagawa, Atsushi
AU - Aso, Yoichi
AU - Suzuki, Mamoru
N1 - Funding Information:
This work was supported in part by a Grant-in-Aid for Scientific Research ( KAKENHI; 21780049 ) from the Ministry of Education, Science, Sports and Culture of Japan and by the Mishima Kaiun Memorial Foundation . This work was also supported in part by the Research Grant for Young Investigators from the Faculty of Agriculture, Kyushu University .
PY - 2012/10
Y1 - 2012/10
N2 - Background: Glutathione transferase (GST) catalyzes glutathione conjugation, a major detoxification pathway for xenobiotics and endogenous substances. Here, we determined the crystal structure of a Delta-class GST from Bombyx mori (bmGSTD) to examine its catalytic residues. Methods: The three-dimensional structure of bmGSTD was resolved by the molecular replacement method and refined to a resolution of 2.0 Å. Results: Structural alignment with a Delta-class GST of Anopheles gambiae indicated that bmGSTD contains 2 distinct domains (an N-terminal domain and a C-terminal domain) connected by a linker. The bound glutathione localized at the N-terminal domain. Putative catalytic residues were changed to alanine by site-directed mutagenesis, and the resulting mutants were characterized in terms of catalytic activity using glutathione and 1-chloro-2,4-dinitrobenzene, a synthetic substrate of GST. Kinetic analysis of bmGSTD mutants indicated that Ser11, Gln51, His52, Ser67, and Arg68 are important for enzyme function. General significance: These results provide structural insights into the catalysis of glutathione conjugation in B. mori by bmGSTD.
AB - Background: Glutathione transferase (GST) catalyzes glutathione conjugation, a major detoxification pathway for xenobiotics and endogenous substances. Here, we determined the crystal structure of a Delta-class GST from Bombyx mori (bmGSTD) to examine its catalytic residues. Methods: The three-dimensional structure of bmGSTD was resolved by the molecular replacement method and refined to a resolution of 2.0 Å. Results: Structural alignment with a Delta-class GST of Anopheles gambiae indicated that bmGSTD contains 2 distinct domains (an N-terminal domain and a C-terminal domain) connected by a linker. The bound glutathione localized at the N-terminal domain. Putative catalytic residues were changed to alanine by site-directed mutagenesis, and the resulting mutants were characterized in terms of catalytic activity using glutathione and 1-chloro-2,4-dinitrobenzene, a synthetic substrate of GST. Kinetic analysis of bmGSTD mutants indicated that Ser11, Gln51, His52, Ser67, and Arg68 are important for enzyme function. General significance: These results provide structural insights into the catalysis of glutathione conjugation in B. mori by bmGSTD.
UR - http://www.scopus.com/inward/record.url?scp=84863193817&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84863193817&partnerID=8YFLogxK
U2 - 10.1016/j.bbagen.2012.04.022
DO - 10.1016/j.bbagen.2012.04.022
M3 - Article
C2 - 22579926
AN - SCOPUS:84863193817
VL - 1820
SP - 1469
EP - 1474
JO - Biochimica et Biophysica Acta - General Subjects
JF - Biochimica et Biophysica Acta - General Subjects
SN - 0304-4165
IS - 10
ER -