Mutagenesis of triad determinants of rat Alox15 alters the specificity of fatty acid and phospholipid oxygenation

Archives of Biochemistry and Biophysics
Mária PekárováDagmar Heydeck

Abstract

Among lipoxygenases ALOX15 orthologs are somewhat peculiar because of their capability of oxygenating polyenoic fatty acids even if they are incorporated in complex lipid-protein assemblies. ALOX15 orthologs of different species have been characterized before, but little is known about the corresponding rat enzyme. Since rats are frequently employed as models in biomedical research we expressed rat Alox15 as recombinant protein in pro- and eukaryotic expression systems and characterized the enzyme with respect to its enzymatic properties. The enzyme oxygenated free arachidonic acid mainly to 12S-HpETE with 15S-HpETE only contributing 10% to the product mixture. Multiple directed mutagenesis studies indicated applicability of the triad concept with particular importance of Leu353 and Ile593 as specificity determinants. Ala404Gly exchange induced subtle alterations in enantioselectivity suggesting partial applicability of the Coffa/Brash concept. Wildtype rat Alox15 and its 15-lipoxygenating Leu353Phe mutant are capable of oxygenating ester lipids of biomembranes and high-density lipoproteins. For the wildtype enzyme 13S-HODE and 12S-HETE were identified as major oxygenation products but for the Leu353Phe mutant 13S-HODE and 15S-...Continue Reading

References

Jul 21, 2004·Trends in Cardiovascular Medicine·Lei Zhao, Colin D Funk

❮ Previous
Next ❯

Citations

Jan 29, 2016·Redox Biology·Fulvio UrsiniHenry Jay Forman
Jul 15, 2016·Proceedings of the National Academy of Sciences of the United States of America·Susan AdelDagmar Heydeck
Oct 21, 2016·Chemphyschem : a European Journal of Chemical Physics and Physical Chemistry·Patricia SauraÀngels González-Lafont
Jul 7, 2020·The FEBS Journal·Florian ReischHartmut Kuhn
Jan 2, 2019·Biochimica Et Biophysica Acta. Molecular and Cell Biology of Lipids·Nikita KozlovHartmut Kuhn
Nov 5, 2019·Biochimica Et Biophysica Acta. Molecular and Cell Biology of Lipids·Marjann SchäferHartmut Kuhn
Mar 11, 2020·Biochimica Et Biophysica Acta. Molecular and Cell Biology of Lipids·Alejandro CruzIgor Ivanov
Nov 26, 2018·Progress in Lipid Research·Nikhlesh K Singh, Gadiparthi N Rao

❮ Previous
Next ❯

Related Concepts

Related Feeds

ApoE, Lipids & Cholesterol

Serum cholesterol, triglycerides, apolipoprotein B (APOB)-containing lipoproteins (very low-density lipoprotein (VLDL), immediate-density lipoprotein (IDL), and low-density lipoprotein (LDL), lipoprotein A (LPA)) and the total cholesterol/high-density lipoprotein (HDL) cholesterol ratio are all connected in diseases. Here is the latest research.