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A topologically conserved aliphatic residue in α-helix 6 stabilizes the hydrophobic core in domain II of glutathione transferases and is a structural determinant for the unfolding pathway

Wallace, Louise A, Blatch, Gregory L and Dirr, Heini W (1998) A topologically conserved aliphatic residue in α-helix 6 stabilizes the hydrophobic core in domain II of glutathione transferases and is a structural determinant for the unfolding pathway. Biochemical Journal, 336 (2). pp. 413-418. ISSN 0264-6021

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Abstract

A topologically conserved residue in a-helix 6 of domain II of human glutathione transferase (hGST) A1-1 was mutated to investigate its contribution to protein stability and the unfolding pathway. The replacement of Leu-164 with alanine (L164A) did not impact on the functional and gross structural properties of native hGST A1-1. The wild-type protein unfolds via a threestate pathway in which only folded dimer and unfolded monomer were highly populated at equilibrium; a native-like dimeric intermediate with partially dissociated domains I and II was detected using stopped-¯ow ¯uorescence studies [Wallace, Sluis- Cremer and Dirr (1998) Biochemistry 37, 5320±5328]. In the present study, urea-induced equilibrium unfolding of L164A hGST A1-1 indicated a destabilization of the native state and suggested the presence of a stable dimeric intermediate. The unfolding kinetic pathway for L164A hGST A1-1, like that for the wild type, is biphasic, with a fast and a slow unfolding event; the cavity-forming mutation has a substantially greater effect on the rate of unfolding of the fast event. The equilibrium and kinetic unfolding data for L164A hGST A1-1 suggest that a rapid pre-equilibrium is established between the native dimer and a dimeric intermediate before complete domain and subunit dissociation and unfolding. It is proposed that the topologically conserved bulky residue in a-helix 6 plays a role in specifying and stabilizing the core of domain II and the interface of domains I and II.

Item Type: Article
Additional Information:

Online ISSN: 1470-8728

Uncontrolled Keywords: ResPubID22235. ∂-helix 6, hGST, protein stability, unfolding pathway
Subjects: Faculty/School/Research Centre/Department > School of Biomedical and Health Sciences
FOR Classification > 0601 Biochemistry and Cell Biology
Depositing User: VUIR
Date Deposited: 13 Dec 2011 01:01
Last Modified: 06 Feb 2015 04:02
URI: http://vuir.vu.edu.au/id/eprint/8180
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Citations in Scopus: 11 - View on Scopus

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