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Publication Detail

Title: Probing lipid-protein adduction with alkynyl surrogates: application to Smith-Lemli-Opitz syndrome.

Authors: Windsor, Katherine; Genaro-Mattos, Thiago C; Kim, Hye-Young H; Liu, Wei; Tallman, Keri A; Miyamoto, Sayuri; Korade, Zeljka; Porter, Ned A

Published In J Lipid Res, (2013 Oct)

Abstract: Lipid modifications aid in regulating (and misregulating) protein function and localization. However, efficient methods to screen for a lipid's ability to modify proteins are not readily available. We present a strategy to identify protein-reactive lipids and apply it to a neurodevelopmental disorder, Smith-Lemli-Opitz syndrome (SLOS). Alkynyl surrogates were synthesized for polyunsaturated fatty acids, phospholipids, cholesterol, 7-dehydrocholesterol (7-DHC), and a 7-DHC-derived oxysterol. To probe for protein-reactive lipids, we used click chemistry to biotinylate the alkynyl tag and detected the lipid-adducted proteins with streptavidin Western blotting. In Neuro2a cells, the trend in amount of protein adduction followed known rates of lipid peroxidation (7-DHC > arachidonic acid > linoleic acid > cholesterol), with alkynyl-7-DHC producing the most adduction among alkynyl lipids. 7-DHC reductase-deficient cells, which cannot properly metabolize 7-DHC, exhibited significantly more alkynyl-7-DHC-protein adduction than control cells. Model studies demonstrated that a 7-DHC peroxidation product covalently modifies proteins. We hypothesize that 7-DHC generates electrophiles that can modify the proteome, contributing to SLOS's complex pathology. These probes and methods would allow for analysis of lipid-modified proteomes in SLOS and other disorders exhibiting 7-DHC accumulation. More broadly, the alkynyl lipid library would facilitate exploration of lipid peroxidation's role in specific biological processes in numerous diseases.

PubMed ID: 23828810 Exiting the NIEHS site

MeSH Terms: Cell Line, Tumor; Cytochromes c/chemistry; Cytochromes c/metabolism; Dehydrocholesterols/chemistry; Dehydrocholesterols/metabolism; Fatty Acids, Unsaturated/chemistry; Fatty Acids, Unsaturated/metabolism; Humans; Lipoylation; Oxidation-Reduction; Phospholipids/chemistry; Phospholipids/metabolism; Protein Processing, Post-Translational*; Proteome/metabolism; Smith-Lemli-Opitz Syndrome/diagnosis; Smith-Lemli-Opitz Syndrome/metabolism*

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