Tertiary interactions stabilise the C-terminal region of human glutathione transferase A1-1: a crystallographic and calorimetric study
dc.contributor.author | Kuhnert, Diane C | |
dc.contributor.author | Sayed, Yasien | |
dc.contributor.author | Mosebi, Salerwe | |
dc.contributor.author | Sayed, Muhammed | |
dc.contributor.author | Sewell, Trevor | |
dc.contributor.author | Dirr, Heini W | |
dc.date.accessioned | 2016-09-01T07:19:51Z | |
dc.date.available | 2016-09-01T07:19:51Z | |
dc.date.issued | 2005 | |
dc.date.updated | 2016-08-30T11:41:13Z | |
dc.description.abstract | The C-terminal region in class Alpha glutathione transferase A1-1 (GSTA1-1), which forms an amphipathic α-helix (helix 9), is known to contribute to the catalytic and non-substrate ligand-binding functions of the enzyme. The region in the apo protein is proposed to be disordered which, upon ligand binding at the active-site, becomes structured and localised. Because Ile219 plays a pivotal role in the stability and localisation of the region, the role of tertiary interactions mediated by Ile219 in determining the conformation and dynamics of the C-terminal region were studied. Ligand-binding microcalorimetric and X-ray structural data were obtained to characterise ligand binding at the active-site and the associated localisation of the C-terminal region. In the crystal structure of the I219A hGSTA1-1·S-hexylglutathione complex, the C-terminal region of one chain is mobile and not observed (unresolved electron density), whereas the corresponding region of the other chain is localised and structured as a result of crystal packing interactions. In solution, the mutant C-terminal region of both chains in the complex is mobile and delocalised resulting in a hydrated, less hydrophobic active-site and a reduction in the affinity of the protein for S-hexylglutathione. Complete dehydration of the active-site, important for maintaining the highly reactive thiolate form of glutathione, requires the binding of ligands and the subsequent localisation of the C-terminal region. Thermodynamic data demonstrate that the mobile C-terminal region in apo hGSTA1-1 is structured and does not undergo ligand-induced folding. Its close proximity to the surface of the wild-type protein is indicated by the concurrence between the observed heat capacity change of complex formation and the type and amount of surface area that becomes buried at the ligand–protein interface when the C-terminal region in the apo protein assumes the same localised structure as that observed in the wild-type complex. | en_ZA |
dc.identifier | http://dx.doi.org/10.1016/j.jmb.2005.04.025 | |
dc.identifier.apacitation | Kuhnert, D. C., Sayed, Y., Mosebi, S., Sayed, M., Sewell, T., & Dirr, H. W. (2005). Tertiary interactions stabilise the C-terminal region of human glutathione transferase A1-1: a crystallographic and calorimetric study. <i>Journal of Molecular Biology</i>, http://hdl.handle.net/11427/21630 | en_ZA |
dc.identifier.chicagocitation | Kuhnert, Diane C, Yasien Sayed, Salerwe Mosebi, Muhammed Sayed, Trevor Sewell, and Heini W Dirr "Tertiary interactions stabilise the C-terminal region of human glutathione transferase A1-1: a crystallographic and calorimetric study." <i>Journal of Molecular Biology</i> (2005) http://hdl.handle.net/11427/21630 | en_ZA |
dc.identifier.citation | Kuhnert, D. C., Sayed, Y., Mosebi, S., Sayed, M., Sewell, T., & Dirr, H. W. (2005). Tertiary interactions stabilise the C-terminal region of human glutathione transferase A1-1: a crystallographic and calorimetric study. Journal of molecular biology, 349(4), 825-838. | en_ZA |
dc.identifier.issn | 0022-2836 | en_ZA |
dc.identifier.ris | TY - Journal Article AU - Kuhnert, Diane C AU - Sayed, Yasien AU - Mosebi, Salerwe AU - Sayed, Muhammed AU - Sewell, Trevor AU - Dirr, Heini W AB - The C-terminal region in class Alpha glutathione transferase A1-1 (GSTA1-1), which forms an amphipathic α-helix (helix 9), is known to contribute to the catalytic and non-substrate ligand-binding functions of the enzyme. The region in the apo protein is proposed to be disordered which, upon ligand binding at the active-site, becomes structured and localised. Because Ile219 plays a pivotal role in the stability and localisation of the region, the role of tertiary interactions mediated by Ile219 in determining the conformation and dynamics of the C-terminal region were studied. Ligand-binding microcalorimetric and X-ray structural data were obtained to characterise ligand binding at the active-site and the associated localisation of the C-terminal region. In the crystal structure of the I219A hGSTA1-1·S-hexylglutathione complex, the C-terminal region of one chain is mobile and not observed (unresolved electron density), whereas the corresponding region of the other chain is localised and structured as a result of crystal packing interactions. In solution, the mutant C-terminal region of both chains in the complex is mobile and delocalised resulting in a hydrated, less hydrophobic active-site and a reduction in the affinity of the protein for S-hexylglutathione. Complete dehydration of the active-site, important for maintaining the highly reactive thiolate form of glutathione, requires the binding of ligands and the subsequent localisation of the C-terminal region. Thermodynamic data demonstrate that the mobile C-terminal region in apo hGSTA1-1 is structured and does not undergo ligand-induced folding. Its close proximity to the surface of the wild-type protein is indicated by the concurrence between the observed heat capacity change of complex formation and the type and amount of surface area that becomes buried at the ligand–protein interface when the C-terminal region in the apo protein assumes the same localised structure as that observed in the wild-type complex. DA - 2005 DB - OpenUCT DP - University of Cape Town J1 - Journal of Molecular Biology LK - https://open.uct.ac.za PB - University of Cape Town PY - 2005 SM - 0022-2836 T1 - Tertiary interactions stabilise the C-terminal region of human glutathione transferase A1-1: a crystallographic and calorimetric study TI - Tertiary interactions stabilise the C-terminal region of human glutathione transferase A1-1: a crystallographic and calorimetric study UR - http://hdl.handle.net/11427/21630 ER - | en_ZA |
dc.identifier.uri | http://hdl.handle.net/11427/21630 | |
dc.identifier.vancouvercitation | Kuhnert DC, Sayed Y, Mosebi S, Sayed M, Sewell T, Dirr HW. Tertiary interactions stabilise the C-terminal region of human glutathione transferase A1-1: a crystallographic and calorimetric study. Journal of Molecular Biology. 2005; http://hdl.handle.net/11427/21630. | en_ZA |
dc.language | eng | en_ZA |
dc.publisher | Elsevier | en_ZA |
dc.publisher.institution | University of Cape Town | |
dc.rights | Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0) | * |
dc.rights.uri | http://creativecommons.org/licenses/by-nc-nd/4.0/ | en_ZA |
dc.source | Journal of Molecular Biology | en_ZA |
dc.source.uri | http://www.journals.elsevier.com/journal-of-molecular-biology/ | |
dc.subject.other | crystallography | |
dc.subject.other | isothermal titration calorimetry | |
dc.subject.other | conformational dynamics | |
dc.subject.other | thermodynamics | |
dc.title | Tertiary interactions stabilise the C-terminal region of human glutathione transferase A1-1: a crystallographic and calorimetric study | en_ZA |
dc.type | Journal Article | en_ZA |
uct.type.filetype | Text | |
uct.type.filetype | Image | |
uct.type.publication | Research | en_ZA |
uct.type.resource | Article | en_ZA |