LDL phospholipid hydrolysis produces modified electronegative particles with an unfolded apoB-100 protein

Asatryan, Liana; Hamilton, Ryan; Isas, J. Mario; Hwang, Juliana; Kayed, Rakez; Sevanian, Alex

doi:10.1194/jlr.m400306-jlr200

Cited by 43 publications

(41 citation statements)

References 41 publications

(51 reference statements)

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“…50 µg of LDL were incubated with 150 µl of LMB cocktail for 1 h to a final volume of 200 µl in 96 well plates. Total tryptophan in apoB-100 was relatively small and any tryptophan hydroperoxides should only be minor contaminants The colorimetric assay was measured at 650 nm after 1 h incubation at room temperature with a leucomethylene blue cocktail mixture [3]. Molar concentrations of lipid peroxides were determined from a standard curve and total yield of LOOH determined as a ratio of µmoles LOOH/g of LDL protein.…”

Section: Lipid Peroxide Measurementsmentioning

confidence: 99%

“…Circular dichroism analysis was used in order to establish an association between specific LDL protein modifications and protein structure [3,11,34]. In vivo LDL sub-fractions ( Fig.…”

Section: Circular Dichroism and Protein Post-translational Modificationsmentioning

confidence: 99%

“…It is widely recognized that oxidative and/or enzyme-mediated modifications of LDL are required for the particle to acquire the inflammatory properties inherent in the initiation and progression of atherosclerosis [3,4]. This notion is strengthened by the observation that posttranslational modifications of apoB100 are elevated in atherosclerotic lesions [5].…”

Section: Introductionmentioning

confidence: 99%

“…Oxidation of LDL can be carried out by transition metals, hemoglobin, myeloperoxidase, cerruloplasmin, and reactive oxygen species generated by vascular endothelium [6][7][8]. The oxidative modifications render the LDL particle electronegatively charged (LDL − ) as compared to native LDL (nLDL) [3,9,10]. Also, LDL − (in vivo oxidatively-modified LDL) contains elevated level of lipid peroxides and aldehydes that are implicated in protein unfolding [11].…”

Section: Introductionmentioning

confidence: 99%

“…Reactive nitrogen species, especially peroxynitrite (ONOO − ), generated by the vascular endothelium, nitrate apo-B-100 in LDL particles [10,[12][13][14][15]. Enzyme-mediated modifications of LDL -accomplished through the action of ubiquitous hydrolytic enzymes-confer atherogenic properties to the lipoprotein particles [4,16]: s-phospholipase A 2 [3] and its free fatty acid product [17], cholesteryl esterases [4], plasmin [18], and matrix metalloproteinase -2 and -9 [18].…”

Section: Introductionmentioning

confidence: 99%

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LDL protein nitration: Implication for LDL protein unfolding

Hamilton

Asatryan

Nilsen

et al. 2008

Archives of Biochemistry and Biophysics

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Oxidatively-or enzymatically-modified low-density lipoprotein (LDL) is intimately involved in the initiation and progression of atherosclerosis. The in vivo modified LDL is electro-negative (LDL − ) and consists of peroxidized lipid and unfolded ApoB-100 protein. This study was aimed at establishing specific protein modifications and conformational changes in LDL − assessed by liquid chromatography/tandem mass spectrometry (LC/MS/MS) and circular dichroism analyses, respectively. The functional significance of these chemical modifications and structural changes were validated with binding and uptake experiments to-and by bovine aortic endothelial cells (BAEC).The plasma LDL − fraction showed increased nitrotyrosine and lipid peroxide content as well as a greater cysteine oxidation as compared with native-and total LDL. LC/MS/MS analyses of LDL − revealed specific modifications in the apoB-100 moiety, largely involving nitration of tyrosines in the α-helical structures and β 2 sheet as well as cysteine oxidation to cysteic acid in β 1 sheet. Circular dichroism analyses showed that the α-helical content of LDL − was substantially lower (~25%) than that of native LDL (~90%); conversely, LDL − showed greater content of β-sheet and random coil structure, in agreement with unfolding of the protein. These results were mimicked by treatment of LDL subfractions with peroxynitrite (ONOO − ) or SIN-1: similar amino acid modifications as well as conformational changes (loss of α-helical structure and gain in β-sheet structure) were observed. Both LDL − and ONOO − -treated LDL showed a statistically significant increase in binding and uptake to-and by BAEC compared to native LDL. We further found that most binding and uptake in control-LDL was through LDL-R with minimal oxLDL-R-dependent uptake. ONOO − -treated LDL was significantly bound and endocytosed by LOX-1, CD36 and SR-A with minimal contribution from LDL-R. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. It is suggested that lipid peroxidation and protein nitration may account for the mechanisms leading to apoB-100 protein unfolding and consequential increase in modified LDL binding and uptake to and by endothelial cells that is dependent on oxLDL scavenger receptors. NIH Public Access

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Section: Lipid Peroxide Measurementsmentioning

confidence: 99%

“…Circular dichroism analysis was used in order to establish an association between specific LDL protein modifications and protein structure [3,11,34]. In vivo LDL sub-fractions ( Fig.…”

Section: Circular Dichroism and Protein Post-translational Modificationsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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LDL protein nitration: Implication for LDL protein unfolding

Hamilton

Asatryan

Nilsen

et al. 2008

Archives of Biochemistry and Biophysics

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Flexible Intravascular EIS Sensors for Detecting Metabolically Active Plaque

Luo

Packard

Abiri

et al. 2020

Interfacing Bioelectronics and Biomedical Sensing

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Amyloid-Forming Properties of Human Apolipoproteins: Sequence Analyses and Structural Insights

Das

Gursky

2015

Advances in Experimental Medicine and Biology

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Apolipoproteins are protein constituents of lipoproteins that transport cholesterol and fat in circulation and are central to cardiovascular health and disease. Soluble apolipoproteins can transiently dissociate from the lipoprotein surface in a labile free form that can misfold, potentially leading to amyloid disease. Misfolding of apoA-I, apoA-II, and serum amyloid A (SAA) causes systemic amyloidoses, apoE4 is a critical risk factor in Alzheimer's disease, and apolipoprotein misfolding is also implicated in cardiovascular disease. To explain why apolipoproteins are overrepresented in amyloidoses, it was proposed that the amphipathic α-helices, which form the lipid surface-binding motif in this protein family, have high amyloid-forming propensity. Here, we use 12 sequence-based bioinformatics approaches to assess amyloid-forming potential of human apolipoproteins and to identify segments that are likely to initiate β-aggregation. Mapping such segments on the available atomic structures of apolipoproteins helps explain why some of them readily form amyloid while others do not. Our analysis shows that nearly all amyloidogenic segments: (i) are largely hydrophobic, (ii) are located in the lipid-binding amphipathic α-helices in the native structures of soluble apolipoproteins, (iii) are predicted in both native α-helices and β-sheets in the insoluble apoB, and (iv) are predicted to form parallel in-register β-sheet in amyloid. Most of these predictions have been verified experimentally for apoC-II, apoA-I, apoA-II and SAA. Surprisingly, the rank order of the amino acid sequence propensity to form amyloid (apoB > apoA-II > apoC-II ≥ apoA-I, apoC-III, SAA, apoC-I > apoA-IV, apoA-V, apoE) does not correlate with the proteins' involvement in amyloidosis. Rather, it correlates directly with the strength of the protein-lipid association, which increases with increasing protein hydrophobicity. Therefore, the lipid surface-binding function and the amyloid-forming propensity are both rooted in apolipoproteins' hydrophobicity, suggesting that functional constraints make it difficult to completely eliminate pathogenic apolipoprotein misfolding. We propose that apolipoproteins have evolved protective mechanisms against misfolding, such as the sequestration of the amyloidogenic segments via the native protein-lipid and protein-protein interactions involving amphipathic α-helices and, in case of apoB, β-sheets. KeywordsLipid transport; Systemic amyloidosis; Atherosclerosis and Alzheimer's disease; Atomic protein structure; Sequence-based prediction algorithms Correspondence to: Olga Gursky. HHS Public AccessAuthor manuscript Adv Exp Med Biol. Author manuscript; available in PMC 2017 September 24.Published in final edited form as:Adv Exp Med Biol. 2015 ; 855: 175-211. doi:10.1007/978-3-319-17344-3_8. Author Manuscript Author ManuscriptAuthor ManuscriptAuthor Manuscript Lipoproteins and Apolipoproteins in Lipid Transport and MetabolismLipids in plasma, lymph and cerebrospinal fluid are transported via lipoproteins that are...

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LDL phospholipid hydrolysis produces modified electronegative particles with an unfolded apoB-100 protein

Cited by 43 publications

References 41 publications

LDL protein nitration: Implication for LDL protein unfolding

LDL protein nitration: Implication for LDL protein unfolding

Flexible Intravascular EIS Sensors for Detecting Metabolically Active Plaque

Amyloid-Forming Properties of Human Apolipoproteins: Sequence Analyses and Structural Insights

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