pH-induced domain interaction and conformational transitions of lipoxygenase-1.

Sudharshan, E and Sonati, Srinivasulu and Appu Rao, A. G. (2000) pH-induced domain interaction and conformational transitions of lipoxygenase-1. Biochimica et Biophysica Acta, 1480. pp. 13-22.

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Abstract

The multidomain structure of soybean LOX1 was examined over the pH range 1-12. Lipoxygenase-1 activity was reversible over broad pH range of 4-10 due to the reversibility of conformational states of the molecule. Below pH 4.0, due to collapse in hydrophobic interactions, the enzyme unfolded to an irreversible conformation with the properties of molten globule state with a mid point of transition at pH 2.4. This intermediate state lost iron irreversibly. In alkaline pH at 11.5, LOX1 underwent partial unfolding with the exposure of cysteine residues with subsequent oxidation of a pair of cysteine residues in the C-terminal domain and this intermediate showed some properties of molten globule state and retained 35% of activity. Beyond pH 12.0, the enzyme was completely inactivated irreversibly due to irreversible conformational changes. The pH-dependent urea-induced unfolding of LOX1 suggested that LOX1 was more stable at pH 7.0 and least stable at pH 9.0. Furthermore, the urea-induced unfolding of LOX1 indicated that the unfolding was biphasic due to pH-dependent domain interactions and involved sequential unfolding of domains. The loss of enzyme activity at pH 4.0 and 7.0 occurred much earlier to unfolding of the C-domain at all pHs studied. The combination of urea-induced unfolding measurements and limited proteolysis experiments suggested that at pH 4.0, the domains in LOX1 were less interactive and existed as tightly folded units. Furthermore, these results confirmed the contribution of ionic interactions in the interdomain contacts.

Item Type:	Article
Additional Information:	Copyright of this article belongs to Elsevier Ltd.
Uncontrolled Keywords:	Lipoxygenase-1; Domain interaction; Molten globule state; Hydrophobic interaction; Unfolding; Limited proteolysis
Subjects:	500 Natural Sciences and Mathematics > 04 Chemistry and Allied Sciences > 29 Protein Chemistry 600 Technology > 08 Food technology > 22 Legumes-Pulses > 05 Soya bean
Divisions:	Protein Chemistry and Technology
Depositing User:	Food Sci. & Technol. Information Services
Date Deposited:	26 Jun 2007 06:22
Last Modified:	17 Oct 2018 10:35
URI:	http://ir.cftri.res.in/id/eprint/1263

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