Molecular basis for the catalytic inactivity of a naturally occurring near-null variant of human ALOX15 |
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| Authors: | Thomas Horn Igor Ivanov Almerinda Di Venere Kumar Reddy Kakularam Pallu Reddanna Melanie L. Conrad Constanze Richter Patrick Scheerer Hartmut Kuhn |
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| Affiliation: | 1. Institute of Biochemistry, University Medicine Berlin-Charité, Charitéplatz 1, D-10117 Berlin, Germany;2. Institute of Toxicology and Pharmacology, University of Rostock, Schillingallee 70, D-18057 Rostock, Germany;3. Department of Experimental Medicine and Surgery, University of Tor Vergata, Via Montpellier 1, I-00133 Rome, Italy;4. School of Life Sciences, University of Hyderabad, Hyderabad 500046, Andhra Pradesh, India;5. National Institute of Animal Biotechnology, Hyderabad 500046, Andhra Pradesh, India;6. Institute of Microbiology and Hygiene, University of Medicine Berlin-Charité, Hindenburgdamm 27, D-12203 Berlin, Germany;7. Institute of Nutrition Technology and Chemistry, Technical University, Gustav-Meyer-Allee, D-13355 Berlin, Germany;8. Institute of Medical physics and Biophysics, University Medicine Berlin-Charité, Charitéplatz 1, D-10117 Berlin, Germany |
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| Abstract: | Mammalian lipoxygenases belong to a family of lipid-peroxidizing enzymes, which have been implicated in cardiovascular, hyperproliferative and neurodegenerative diseases. Here we report that a naturally occurring mutation in the hALOX15 gene leads to expression of a catalytically near-null enzyme variant (hGly422Glu). The inactivity may be related to severe misfolding of the enzyme protein, which was concluded from CD-spectra as well as from thermal and chemical stability assays. In silico mutagenesis experiments suggest that most mutations at hGly422 have the potential to induce sterical clash, which might be considered a reason for protein misfolding. hGly422 is conserved among ALOX5, ALOX12 and ALOX15 isoforms and corresponding hALOX12 and hALOX5 mutants also exhibited a reduced catalytic activity. Interestingly, in the hALOX5 Gly429Glu mutants the reaction specificity of arachidonic acid oxygenation was shifted from 5S- to 8S- and 12R-H(p)ETE formation. Taken together, our data indicate that the conserved glycine is of functional importance for these enzyme variants and most mutants at this position lose catalytic activity. |
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