title: Structural basis of the suppressed catalytic activity of wild-type human glutathione transferase T1-1 compared to its W234R mutant.
abstract: The crystal structures of wild-type human theta class glutathione-S-transferase (GST) T1-1 and its W234R mutant, where Trp234 was replaced by Arg, were solved both in the presence and absence of S-hexyl-glutathione. The W234R mutant was of interest due to its previously observed enhanced catalytic activity compared to the wild-type enzyme. GST T1-1 from rat and mouse naturally contain Arg in position 234, with correspondingly high catalytic efficiency. The overall structure of GST T1-1 is similar to that of GST T2-2, as expected from their 53% sequence identity at the protein level. Wild-type GST T1-1 has the side-chain of Trp234 occupying a significant portion of the active site. This bulky residue prevents efficient binding of both glutathione and hydrophobic substrates through steric hindrance. The wild-type GST T1-1 crystal structure, obtained from co-crystallization experiments with glutathione and its derivatives, showed no electron density for the glutathione ligand. However, the structure of GST T1-1 mutant W234R showed clear electron density for S-hexyl-glutathione after co-crystallization. In contrast to Trp234 in the wild-type structure, the side-chain of Arg234 in the mutant does not occupy any part of the substrate-binding site. Instead, Arg234 is pointing in a different direction and, in addition, interacts with the carboxylate group of glutathione. These findings explain our earlier observation that the W234R mutant has a markedly improved catalytic activity with most substrates tested to date compared to the wild-type enzyme. GST T1-1 catalyzes detoxication reactions as well as reactions that result in toxic products, and our findings therefore suggest that humans have gained an evolutionary advantage by a partially disabled active site.
authors.affiliation: Department of Cell and Molecular Biology, Uppsala University, Biomedical Centre, Box 590, SE-751 24, Uppsala, Sweden. kaspars@xray.bmc.uu.se
authors.forename: Kaspars
authors.initials: K
authors.lastname: Tars
authors.forename: Anna-Karin
authors.initials: AK
authors.lastname: Larsson
authors.forename: Abeer
authors.initials: A
authors.lastname: Shokeer
authors.forename: Birgit
authors.initials: B
authors.lastname: Olin
authors.forename: Bengt
authors.initials: B
authors.lastname: Mannervik
authors.forename: Gerard J
authors.initials: GJ
authors.lastname: Kleywegt
mesh_terms: D001665:Binding Sites / Q000235:genetics; D002384:Catalysis; D018360:Crystallography, X-Ray; D005978:Glutathione / Q000031:analogs & derivatives / Q000737:chemistry; D005982:Glutathione Transferase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D020125:Mutation, Missense; D011487:Protein Conformation
chemical_list: C491107:GSTT2 protein, human; D005982:Glutathione Transferase; C495790:glutathione transferase T1-1, human; D005978:Glutathione; C033550:hexylglutathione