Reagent or myeloperoxidase-generated hypochlorite affects discrete regions in lipid-free and lipid-associated human apolipoprotein A-I

115

To determine whether oxidative modification of apoA-I tyrosine residues was responsible for the MPO-mediated inactivation of cholesterol acceptor activity, we made recombinant apoAI with site-specific substitutions of all seven tyrosine residues to phenylalanine. ApoAI and the tyrosine-free apoAI were equally susceptible to dosedependent MPO-mediated loss of ABCA1-dependent cholesterol acceptor activity, as well as lipid binding activity. MPO modification altered the migration of apoAI on SDS gels and decreased its ␣-helix content. MPO-induced modification also targeted apoAI tryptophan and lysine residues. Specifically, we detected apoAI tryptophan oxidation to mono-and dihydroxytryptophan and apoAI lysine modification to chlorolysine and 2-aminoadipic acid. Thus, tyrosine modification of apoAI is not required for its MPO-mediated inhibition of cholesterol acceptor activity.The levels of high density lipoprotein (HDL) 2 and its major protein component, apolipoprotein AI (apoAI), are inversely correlated to the risk of coronary artery disease in developed countries (1). Their protective effects may be mediated by several activities, including their ability to remove excess cholesterol from peripheral tissues for delivery to the liver via the reverse cholesterol transport pathway. Cellular lipid efflux to apoAI, as well as HDL biogenesis, is mediated by the transporter ABCA1, which is defective in Tangier disease (2-5). Although apoAI may have additional anti-atherogenic properties, recent studies in the mouse have proved that apoAI can stimulate the removal of the preloaded cholesterol from macrophages and the appearance of this cholesterol in the feces (6). ApoAI is a 243-residue polypeptide containing an N-terminal domain followed by ten 11-or 22-residue amphipathic ␣-helical domains (7). ApoAI adopts two distinct physiological conformations, lipid-free and lipid-bound. In the absence of lipid, apoAI is thought to exist as a helical bundle or hairpin that is stabilized by hydrophobic helix-helix interactions (8 -12). These helical bundles undergo a conformational change during lipid association that results in the substitution of helix-helix interactions for helix-lipid interactions (13).Myeloperoxidase (MPO) is a neutrophil and monocyte/macrophage enzyme that utilizes hydrogen peroxide and chloride to generate the chlorinating oxidant HOCl to kill pathogens (14). MPO can also utilize hydrogen peroxide and nitrite to generate nitrating oxidants capable of initiating lipid peroxidation and protein nitration (15). MPO can promote both protein nitration and lipid peroxidation in vivo (16 -18), and the blood levels of MPO and nitrotyrosine are predictors of an increased risk of cardiovascular disease (19 -21). We recently described that plasma and atherosclerotic lesion-derived apoAI are selective targets for MPO modification resulting in the appearance of specific nitro-and chlorotyrosine residues, and that this modification dose-dependently decreased the cholesterol acceptor and lipid binding activities of apoAI...

Section: Mpo-induced Changes In Apoai Gelsupporting

confidence: 72%

mentioning

confidence: 99%

Tyrosine Modification Is Not Required for Myeloperoxidase-induced Loss of Apolipoprotein A-I Functional Activities

Peng

Brubaker

et al. 2005

115

“…This contrasts with the oneelectron oxidation that produces core aldehydes via ␤-scission of the fatty acyl alkoxyl radical (46). Oxidation-sensitive amino acids that may be involved would include tyrosine, which is well known to form tyrosyl free radical (7,47), tryptophan, and histidine (48). Tyrosyl free radical has been shown to modify apoA-I and apoA-II and promote apoA-I:apoA-II dimer formation in HDL (7) and dityrosine formation in LDL (47).…”

Section: Discussionmentioning

confidence: 96%

Apolipoprotein A-I Promotes the Formation of Phosphatidylcholine Core Aldehydes That Are Hydrolyzed by Paraoxonase (PON-1) during High Density Lipoprotein Oxidation with a Peroxynitrite Donor

Ahmed

Ravandi²,

Maguire

et al. 2001

133

High density lipoprotein (HDL) is rich in polyunsaturated phospholipids that are sensitive to oxidation. However, the effect of apolipoprotein A-I and paraoxonase-1 (PON-1) on phosphatidylcholine oxidation products has not been identified. We subjected native HDL, trypsinized HDL, and HDL lipid suspensions to oxidation by the peroxynitrite donor, 3-morpholinosydnonimine. HDL had a basal level of phosphatidylcholine mono-and di-hydroperoxides that increased to a greater extent in HDL, compared with either trypsinized HDL or HDL lipid alone. Phosphatidylcholine core aldehydes, which were present in small amounts, increased 10-fold during oxidation of native HDL, compared with trypsinized HDL (p ‫؍‬ 0.004), and 4-fold compared with HDL lipid suspensions (p ‫؍‬ 0.0021). In addition, the content of lysophosphatidylcholine increased 300% during oxidation of native HDL, but only 80 and 25%, respectively, during oxidation of trypsinized HDL and HDL lipid suspensions. Phosphatidylcholine isoprostanes accumulated in comparable amounts during the oxidation of all three preparations. Incubation of apolipoprotein A-I with 1-palmitoyl-2-linoleoyl glycerophosphocholine proteoliposomes in the presence of 3-morpholinosydnonimine or apoAI with phosphatidylcholine hydroperoxides resulted in a significant increase in phosphatidylcholine core aldehydes with no formation of lysophosphatidylcholine. We propose that apolipoprotein A-I catalyzes a one-electron oxidation of alkoxyl radicals. Purified PON-1 hydrolyzed phosphatidylcholine core aldehydes to lysophosphatidylcholine. We conclude that, upon HDL oxidation with peroxynitrite, apolipoprotein AI increases the formation of phosphatidylcholine core aldehydes that are subsequently hydrolyzed by PON1.

“…Lipoproteins recovered by direct aspiration were desalted by size-exclusion chromatography on PD-10 columns, and purity was confirmed via SDS-PAGE identification of associated proteins. ApoA-I was isolated as described (28).…”

Section: Methodsmentioning

confidence: 99%

ABCA1 and Scavenger Receptor Class B, Type I, Are Modulators of Reverse Sterol Transport at an in Vitro Blood-Brain Barrier Constituted of Porcine Brain Capillary Endothelial Cells

Panzenboeck

Balázs

Sovic

et al. 2002

Self Cite

185

151