Human 15-LOX-1 active site mutations alter inhibitor binding and decrease potency. Issue 21 (1st November 2016)

• Record Type: Journal Article
• Authors: Armstrong, Michelle; van Hoorebeke, Christopher; Horn, Thomas; Deschamps, Joshua; Freedman, J. Cody; Kalyanaraman, Chakrapani; Jacobson, Matthew P.; Holman, Theodore
• Is Part Of: Bioorganic & medicinal chemistry. Volume 24:Issue 21(2016)
• Page Start: 5380
• Page End: 5387
• Publication Date: 2016-11-01
• Subjects: LOX lipoxygenase -- h15-LOX-1 human reticulocyte 15-lipoxygenase-1 -- tPA tissue plasminogen activator -- FDA Food and Drug Administration -- s15-LOX-1 soybean 15-lipoxygenase-1 -- r15-LOX rabbit reticulocyte 15-LOX -- AA arachidonic acid -- 12-HpETE 12-hydroperoxyeicosatetraenoic acid -- LA linoleic acid -- 9R-HpODE 9-(R)-hydroperoxyoctadecadienoic acid -- 13-HpODE 13-(S)-hydroperoxyoctadecadienoic acid -- 15-HpETE 15-hydroperoxyeicosatetraenoic acid -- ICP-MS inductively coupled plasma mass spectrometer -- EDTA ethylenediaminetetraacetic acid -- BSA bovine serum albumin -- SDS–PAGE sodium dodecyl sulfate–poly-acrylamide gel electrophoresis -- RMSD root-mean-squared-distance; Human -- 15-Lipoxygenase-1 -- Mutagenesis -- Inhibitor -- Docking; Bioorganic chemistry -- Periodicals; Pharmaceutical chemistry -- Periodicals; Biochemistry -- Periodicals; Chemistry, Clinical -- Periodicals; Chemistry, Organic -- Periodicals; Chimie bio-organique -- Périodiques; Chimie pharmaceutique -- Périodiques; 615.19
• DOI: 10.1016/j.bmc.2016.08.063
• Languages: English
• ISSNs: 0968-0896

Abstract

Graphical abstract: Abstract: Human 15-lipoxygenase-1 (h15-LOX-1 or h12/15-LOX) reacts with polyunsaturated fatty acids and produces bioactive lipid derivatives that are implicated in many important human diseases. One such disease is stroke, which is the fifth leading cause of death and the first leading cause of disability in America. The discovery of h15-LOX-1 inhibitors could potentially lead to novel therapeutics in the treatment of stroke, however, little is known about the inhibitor/active site interaction. This study utilizes site-directed mutagenesis, guided in part by molecular modeling, to gain a better structural understanding of inhibitor interactions within the active site. We have generated eight mutants (R402L, R404L, F414I, F414W, E356Q, Q547L, L407A, I417A) of h15-LOX-1 to determine whether these active site residues interact with two h15-LOX-1 inhibitors, ML351 and anML094 derivative, compound18 . IC50 values and steady-state inhibition kinetics were determined for the eight mutants, with four of the mutants affecting inhibitor potency relative to wild type h15-LOX-1 (F414I, F414W, E356Q and L407A). The data indicate thatML351 and compound18, bind in a similar manner in the active site to an aromatic pocket close to F414 but have subtle differences in their specific binding modes. This information establishes the binding mode forML094 andML351 and will be leveraged to develop next-generation inhibitors.