Identification and characterization of a new protein isoform of human 5-lipoxygenase

  • Leukotrienes (LTs) are inflammatory mediators that play a pivotal role in many diseases like asthma bronchiale, atherosclerosis and in various types of cancer. The key enzyme for generation of LTs is the 5-lipoxygenase (5-LO). Here, we present a novel putative protein isoform of human 5-LO that lacks exon 4, termed 5-LOΔ4, identified in cells of lymphoid origin, namely the Burkitt lymphoma cell lines Raji and BL41 as well as primary B and T cells. Deletion of exon 4 does not shift the reading frame and therefore the mRNA is not subjected to non-mediated mRNA decay (NMD). By eliminating exon 4, the amino acids Trp144 until Ala184 are omitted in the corresponding protein. Transfection of HEK293T cells with a 5-LOΔ4 expression plasmid led to expression of the corresponding protein which suggests that the 5-LOΔ4 isoform is a stable protein in eukaryotic cells. We were also able to obtain soluble protein after expression in E. coli and purification. The isoform itself lacks canonical enzymatic activity as it misses the non-heme iron but it still retains ATP-binding affinity. Differential scanning fluorimetric analysis shows two transitions, corresponding to the two domains of 5-LO. Whilst the catalytic domain of 5-LO WT is destabilized by calcium, addition of calcium has no influence on the catalytic domain of 5-LOΔ4. Furthermore, we investigated the influence of 5-LOΔ4 on the activity of 5-LO WT and proved that it stimulates 5-LO product formation at low protein concentrations. Therefore regulation of 5-LO by its isoform 5-LOΔ4 might represent a novel mechanism of controlling the biosynthesis of lipid mediators.

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Metadaten
Author:Ann-Kathrin Häfner, Kim Beilstein, Philipp Graab, Ann-Katrin Ball, Meike J. Saul, Bettina Hofmann, Dieter SteinhilberORCiDGND
URN:urn:nbn:de:hebis:30:3-419916
DOI:https://doi.org/10.1371/journal.pone.0166591
ISSN:1932-6203
Pubmed Id:https://pubmed.ncbi.nlm.nih.gov/27855198
Parent Title (English):PLoS One
Publisher:PLoS
Place of publication:Lawrence, Kan.
Document Type:Article
Language:English
Date of Publication (online):2016/11/21
Date of first Publication:2016/11/17
Publishing Institution:Universitätsbibliothek Johann Christian Senckenberg
Release Date:2016/11/21
Volume:11
Issue:(11):e0166591
Page Number:16
HeBIS-PPN:402528794
Institutes:Biochemie, Chemie und Pharmazie / Pharmazie
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie
Sammlungen:Universitätspublikationen
Sammlung Biologie / Sondersammelgebiets-Volltexte
Open-Access-Publikationsfonds:Biochemie, Chemie und Pharmazie
Licence (German):License LogoCreative Commons - Namensnennung 4.0