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A mutation interfering with 5-lipoxygenase domain interaction leads to increased enzyme activity
Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Computational and Systems Biology.
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2014 (English)In: Archives of Biochemistry and Biophysics, ISSN 0003-9861, E-ISSN 1096-0384, Vol. 545, 179-185 p.Article in journal (Refereed) Published
Abstract [en]

5-Lipoxygenase (5-LOX) catalyzes two steps in conversion of arachidonic acid to proinflammatory leukotrienes. Lipoxygenases, including human 5-LOX, consist of an N-terminal C2-like beta-sandwich and a catalytic domain. We expressed the 5-LOX domains separately, these were found to interact in the yeast two-hybrid system. The 5-LOX structure suggested association between Arg(101) in the beta-sandwich and Asp(166) in the catalytic domain, due to electrostatic interaction as well as hydrogen bonds. Indeed, mutagenic replacements of these residues led to loss of two-hybrid interaction. Interestingly, when Arg(101) was mutated to Asp in intact 5-LOX, enzyme activity was increased. Thus, higher initial velocity of the reaction (v(init)) and increased final amount of products were monitored for 5-LOX-R101D, at several different assay conditions. In the 5-LOX crystal structure, helix alpha 2 and adjacent loops (including Asp(166)) of the 5-LOX catalytic domain has been proposed to form a flexible lid controlling access to the active site, and lid movement would be determined by bonding of lid residues to the C2-like beta-sandwich. The more efficient catalysis following disruption of the R101-D166 ionic association supports the concept of such a flexible lid in human 5-LOX. (C) 2014 Elsevier Inc. All rights reserved.

Place, publisher, year, edition, pages
2014. Vol. 545, 179-185 p.
Keyword [en]
Leukotriene, Eicosanoid, Arachidonic acid, Two-hybrid system, Domain interaction
National Category
Biological Sciences
URN: urn:nbn:se:uu:diva-222931DOI: 10.1016/j.abb.2014.01.017ISI: 000332751700022OAI: oai:DiVA.org:uu-222931DiVA: diva2:713030
Available from: 2014-04-17 Created: 2014-04-15 Last updated: 2014-04-17Bibliographically approved

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Persson, Bengt
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