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Structural insights into the dehydroascorbate reductase activity of human omega-class glutathione transferases

Journal Article


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Abstract


  • The reduction of dehydroascorbate (DHA) to ascorbic acid (AA) is a vital cellular function. The omega-class glutathione transferases (GSTs) catalyze several reductive reactions in cellular biochemistry, including DHA reduction. In humans, two isozymes (GSTO1-1 and GSTO2-2) with significant DHA reductase (DHAR) activity are found, sharing 64% sequence identity. While the activity of GSTO2-2 is higher, it is significantly more unstable in vitro. We report the first crystal structures of human GSTO2-2, stabilized through site-directed mutagenesis and determined at 1.9 Ã� resolution in the presence and absence of glutathione (GSH). The structure of a human GSTO1-1 has been determined at 1.7 Ã� resolution in complex with the reaction product AA, which unexpectedly binds in the G-site, where the glutamyl moiety of GSH binds. The structure suggests a similar mode of ascorbate binding in GSTO2-2. This is the first time that a non-GSH-based reaction product has been observed in the G-site of any GST. AA stacks against a conserved aromatic residue, F34 (equivalent to Y34 in GSTO2-2). Mutation of Y34 to alanine in GSTO2-2 eliminates DHAR activity. From these structures and other biochemical data, we propose a mechanism of substrate binding and catalysis of DHAR activity.

Authors


  •   Zhou, Huina (external author)
  •   Brock, Joseph (external author)
  •   Liu, Dan (external author)
  •   Board, Philip G. (external author)
  •   Oakley, Aaron J.

Publication Date


  • 2012

Citation


  • Zhou, H., Brock, J., Liu, D., Board, P. G. & Oakley, A. J. (2012). Structural insights into the dehydroascorbate reductase activity of human omega-class glutathione transferases. Journal of Molecular Biology, 420 (3), 190-203.

Scopus Eid


  • 2-s2.0-84862022469

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=8103&context=scipapers

Ro Metadata Url


  • http://ro.uow.edu.au/scipapers/4760

Number Of Pages


  • 13

Start Page


  • 190

End Page


  • 203

Volume


  • 420

Issue


  • 3

Abstract


  • The reduction of dehydroascorbate (DHA) to ascorbic acid (AA) is a vital cellular function. The omega-class glutathione transferases (GSTs) catalyze several reductive reactions in cellular biochemistry, including DHA reduction. In humans, two isozymes (GSTO1-1 and GSTO2-2) with significant DHA reductase (DHAR) activity are found, sharing 64% sequence identity. While the activity of GSTO2-2 is higher, it is significantly more unstable in vitro. We report the first crystal structures of human GSTO2-2, stabilized through site-directed mutagenesis and determined at 1.9 Ã� resolution in the presence and absence of glutathione (GSH). The structure of a human GSTO1-1 has been determined at 1.7 Ã� resolution in complex with the reaction product AA, which unexpectedly binds in the G-site, where the glutamyl moiety of GSH binds. The structure suggests a similar mode of ascorbate binding in GSTO2-2. This is the first time that a non-GSH-based reaction product has been observed in the G-site of any GST. AA stacks against a conserved aromatic residue, F34 (equivalent to Y34 in GSTO2-2). Mutation of Y34 to alanine in GSTO2-2 eliminates DHAR activity. From these structures and other biochemical data, we propose a mechanism of substrate binding and catalysis of DHAR activity.

Authors


  •   Zhou, Huina (external author)
  •   Brock, Joseph (external author)
  •   Liu, Dan (external author)
  •   Board, Philip G. (external author)
  •   Oakley, Aaron J.

Publication Date


  • 2012

Citation


  • Zhou, H., Brock, J., Liu, D., Board, P. G. & Oakley, A. J. (2012). Structural insights into the dehydroascorbate reductase activity of human omega-class glutathione transferases. Journal of Molecular Biology, 420 (3), 190-203.

Scopus Eid


  • 2-s2.0-84862022469

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=8103&context=scipapers

Ro Metadata Url


  • http://ro.uow.edu.au/scipapers/4760

Number Of Pages


  • 13

Start Page


  • 190

End Page


  • 203

Volume


  • 420

Issue


  • 3