Probes for protein adduction in cholesterol biosynthesis disorders: Alkynyl lanosterol as a viable sterol precursor
The formation of lipid electrophile-protein adducts is associated with many disorders that involve perturbations of cellular redox status. The identities of adducted proteins and the effects of adduction on protein function are mostly unknown and an increased understanding of these factors may help...
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| Format: | Article |
| Language: | English |
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Elsevier
2017-08-01
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| Series: | Redox Biology |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2213231717300216 |
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| author | Keri A. Tallman Hye-Young H. Kim Zeljka Korade Thiago C. Genaro-Mattos Phillip A. Wages Wei Liu Ned A. Porter |
| author_facet | Keri A. Tallman Hye-Young H. Kim Zeljka Korade Thiago C. Genaro-Mattos Phillip A. Wages Wei Liu Ned A. Porter |
| author_sort | Keri A. Tallman |
| collection | DOAJ |
| description | The formation of lipid electrophile-protein adducts is associated with many disorders that involve perturbations of cellular redox status. The identities of adducted proteins and the effects of adduction on protein function are mostly unknown and an increased understanding of these factors may help to define the pathogenesis of various human disorders involving oxidative stress. 7-Dehydrocholesterol (7-DHC), the immediate biosynthetic precursor to cholesterol, is highly oxidizable and gives electrophilic oxysterols that adduct proteins readily, a sequence of events proposed to occur in Smith-Lemli-Opitz syndrome (SLOS), a human disorder resulting from an error in cholesterol biosynthesis. Alkynyl lanosterol (a-Lan) was synthesized and studied in Neuro2a cells, Dhcr7-deficient Neuro2a cells and human fibroblasts. When incubated in control Neuro2a cells and control human fibroblasts, a-Lan completed the sequence of steps involved in cholesterol biosynthesis and alkynyl-cholesterol (a-Chol) was the major product formed. In Dhcr7-deficient Neuro2a cells or fibroblasts from SLOS patients, the biosynthetic transformation was interrupted at the penultimate step and alkynyl-7-DHC (a-7-DHC) was the major product formed. When a-Lan was incubated in Dhcr7-deficient Neuro2a cells and the alkynyl tag was used to ligate a biotin group to alkyne-containing products, protein-sterol adducts were isolated and identified. In parallel experiments with a-Lan and a-7-DHC in Dhcr7-deficient Neuro2a cells, a-7-DHC was found to adduct to a larger set of proteins (799) than a-Lan (457) with most of the a-Lan protein adducts (423) being common to the larger a-7-DHC set. Of the 423 proteins found common to both experiments, those formed from a-7-DHC were more highly enriched compared to a DMSO control than were those derived from a-Lan. The 423 common proteins were ranked according to the enrichment determined for each protein in the a-Lan and a-7-DHC experiments and there was a very strong correlation of protein ranks for the adducts formed in the parallel experiments. Keywords: Alkynyl sterols, 7-dehydrocholesterol, Cholesterol, HPLC-MS, GC-MS, DHCR7, Smith-Lemli-Opitz Syndrome |
| first_indexed | 2024-12-11T02:32:37Z |
| format | Article |
| id | doaj.art-9d341d16b651429fb5861430ed2a297a |
| institution | Directory Open Access Journal |
| issn | 2213-2317 |
| language | English |
| last_indexed | 2024-12-11T02:32:37Z |
| publishDate | 2017-08-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Redox Biology |
| spelling | doaj.art-9d341d16b651429fb5861430ed2a297a2022-12-22T01:23:48ZengElsevierRedox Biology2213-23172017-08-0112182190Probes for protein adduction in cholesterol biosynthesis disorders: Alkynyl lanosterol as a viable sterol precursorKeri A. Tallman0Hye-Young H. Kim1Zeljka Korade2Thiago C. Genaro-Mattos3Phillip A. Wages4Wei Liu5Ned A. Porter6Department of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235, United StatesDepartment of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235, United StatesVanderbilt Kennedy Center for Research on Human Development, Vanderbilt University, Nashville, TN 37235, United States; Department of Psychiatry, Vanderbilt University, Nashville, TN 37235, United StatesDepartment of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235, United StatesDepartment of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235, United StatesDepartment of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235, United StatesDepartment of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235, United States; Vanderbilt Kennedy Center for Research on Human Development, Vanderbilt University, Nashville, TN 37235, United States; Corresponding author at: Department of Chemistry and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, TN 37235, United States.The formation of lipid electrophile-protein adducts is associated with many disorders that involve perturbations of cellular redox status. The identities of adducted proteins and the effects of adduction on protein function are mostly unknown and an increased understanding of these factors may help to define the pathogenesis of various human disorders involving oxidative stress. 7-Dehydrocholesterol (7-DHC), the immediate biosynthetic precursor to cholesterol, is highly oxidizable and gives electrophilic oxysterols that adduct proteins readily, a sequence of events proposed to occur in Smith-Lemli-Opitz syndrome (SLOS), a human disorder resulting from an error in cholesterol biosynthesis. Alkynyl lanosterol (a-Lan) was synthesized and studied in Neuro2a cells, Dhcr7-deficient Neuro2a cells and human fibroblasts. When incubated in control Neuro2a cells and control human fibroblasts, a-Lan completed the sequence of steps involved in cholesterol biosynthesis and alkynyl-cholesterol (a-Chol) was the major product formed. In Dhcr7-deficient Neuro2a cells or fibroblasts from SLOS patients, the biosynthetic transformation was interrupted at the penultimate step and alkynyl-7-DHC (a-7-DHC) was the major product formed. When a-Lan was incubated in Dhcr7-deficient Neuro2a cells and the alkynyl tag was used to ligate a biotin group to alkyne-containing products, protein-sterol adducts were isolated and identified. In parallel experiments with a-Lan and a-7-DHC in Dhcr7-deficient Neuro2a cells, a-7-DHC was found to adduct to a larger set of proteins (799) than a-Lan (457) with most of the a-Lan protein adducts (423) being common to the larger a-7-DHC set. Of the 423 proteins found common to both experiments, those formed from a-7-DHC were more highly enriched compared to a DMSO control than were those derived from a-Lan. The 423 common proteins were ranked according to the enrichment determined for each protein in the a-Lan and a-7-DHC experiments and there was a very strong correlation of protein ranks for the adducts formed in the parallel experiments. Keywords: Alkynyl sterols, 7-dehydrocholesterol, Cholesterol, HPLC-MS, GC-MS, DHCR7, Smith-Lemli-Opitz Syndromehttp://www.sciencedirect.com/science/article/pii/S2213231717300216 |
| spellingShingle | Keri A. Tallman Hye-Young H. Kim Zeljka Korade Thiago C. Genaro-Mattos Phillip A. Wages Wei Liu Ned A. Porter Probes for protein adduction in cholesterol biosynthesis disorders: Alkynyl lanosterol as a viable sterol precursor Redox Biology |
| title | Probes for protein adduction in cholesterol biosynthesis disorders: Alkynyl lanosterol as a viable sterol precursor |
| title_full | Probes for protein adduction in cholesterol biosynthesis disorders: Alkynyl lanosterol as a viable sterol precursor |
| title_fullStr | Probes for protein adduction in cholesterol biosynthesis disorders: Alkynyl lanosterol as a viable sterol precursor |
| title_full_unstemmed | Probes for protein adduction in cholesterol biosynthesis disorders: Alkynyl lanosterol as a viable sterol precursor |
| title_short | Probes for protein adduction in cholesterol biosynthesis disorders: Alkynyl lanosterol as a viable sterol precursor |
| title_sort | probes for protein adduction in cholesterol biosynthesis disorders alkynyl lanosterol as a viable sterol precursor |
| url | http://www.sciencedirect.com/science/article/pii/S2213231717300216 |
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