Helical remodeling augments 5-lipoxygenase activity in the synthesis of proinflammatory mediators

Authors

Eden M. Gallegos, LSU Health Sciences Center - New OrleansFollow
Tanner D. Reed, LSU Health Sciences Center - New OrleansFollow
Forge A. Mathes, Louisiana State University, Baton Rouge, LA
Nelson V. Guevara, Louisiana State University, Baton Rouge, LA
David B. Neau, Argonne National Laboratory, Argonne, IL
Wei Huang, Case Western Reserve University, Cleveland, OH
Marcia E. Newcomer, Louisiana State University, Baton Rouge, LA
Nathaniel C. Gilbert, Louisiana State University, Baton Rouge, LA

Document Type

Article

Publication Date

9-1-2022

Publication Title

Journal of Biological Chemistry

Abstract

The synthesis of proinflammatory leukotrienes implicated in asthma, allergic rhinitis, and atherosclerosis is initiated by the enzyme 5-lipoxygenase (5-LOX). The crystal structure of human Stable-5-LOX revealed a conformation where the catalytic iron was inaccessible to bulk solvent as two aromatic residues on a conserved helix-α2 (Hα2) plugged the substrate access portal. Whether 5-LOX can also adopt a more open conformation has not been resolved. Here, we present a new conformation of 5-LOX where Hα2 adopts an elongated conformation equivalent to that described in other animal lipoxygenase structures. Our observation of the sigmoidal kinetic behavior of 5-LOX, which is indicative of positive cooperativity, is consistent with a substrate-induced conformational change that shifts the ensemble of enzyme populations to favor the catalytically competent state. Strategic point mutations along Hα2 designed to unlock the closed conformation and elongate Hα2 resulted in improved kinetic parameters, altered limited proteolysis data, and a drastic reduction in the length of the lag phase yielding the most active Stable-5-LOX to date. Structural predictions by AlphaFold2 of these variants statistically favor an elongated Hα2 and reinforce a model in which improved kinetic parameters correlate with a more readily adopted open conformation. Taken together, these data provide valuable insights into the synthesis of leukotrienes.

First Page

1

Last Page

14

Issue

9

Publisher

Elsevier

Creative Commons License

This work is licensed under a Creative Commons Attribution-NonCommercial-No Derivative Works 4.0 International License.

Recommended Citation

Gallegos, Eden M.; Reed, Tanner D.; Mathes, Forge A.; Guevara, Nelson V.; Neau, David B.; Huang, Wei; Newcomer, Marcia E.; and Gilbert, Nathaniel C., "Helical remodeling augments 5-lipoxygenase activity in the synthesis of proinflammatory mediators" (2022). School of Medicine Faculty Publications. 4445.
https://digitalscholar.lsuhsc.edu/som_facpubs/4445
10.1016/j.jbc.2022.102282