The Lipid Peroxidation Product, 4‐Hydroxy‐2‐trans‐Nonenal, Alters the Conformation of Cortical Synaptosomal Membrane Proteins

Abstract

Alzheimer's disease (AD) is widely held to be a disorder associated with oxidative stress due, in part, to the membrane action of amyloid~3-peptide (A/3). A~-associated free radicals cause lipid peroxidation, a major product of which is 4-hydroxy-2-trans-nonenal (HNE). We determined whether HNE would alter the conformation of synaptosomal membrane proteins, which might be related to the known neurotoxicity of A~3and HNE. Electron paramagnetic resonance spectroscopy, using a protein-specific spin label, MAL-6 (2,2 ,6,6-tetramethyl-4-maleimidopiperidin-1-oxyl), was used to probe conformational changes in gerbil cortical synaptosomal membrane proteins, and a lipid-specific stearic acid label, 5-nitroxide stearate, was used to probe for HNE-induced alterations in the fluidity of the bilayer domain of these membranes. Synaptosomal membranes, incubated with low concentrations of HNE, exhibited changes in protein conformation and bilayer order and motion (fluidity). The changes in protein conformation were found to be concentration-and time-dependent. Significant protein conformational changes were observed at physiologically relevant concentrations of 1-10 p~MHNE, reminiscent of similar changes in synaptosomal membrane proteins from senile plaque-and A/3-rich AD hippocampal and inferior parietal brain regions. HNE-induced modifications in the physical state of gerbil synaptosomal membrane proteins were prevented completely by using excess glutathione ethyl ester, known to protect neurons from HNEcaused neurotoxicity. Membrane fluidity was found to increase at higher concentrations of HNE (50 /2M). The results obtained are discussed with relevance to the hypothesis of A/3-induced free radical-mediated lipid peroxidation, leading to subsequent HNE-induced alterations in the structure and function of key membrane proteins with consequent neurotoxicity in AD brain.

Contribution of Lipid Dynamics on the Inhibition of Bovine Brain Synaptosomal Na+-K+-ATPase Activity Induced by 4-Hydroxy-2-nonenal

“…24) From these findings, therefore, it is suggested that the lipid fluidity at the membrane surface decreased by modification of the synaptosomes with HNE. This finding is not consistent with the reports by Subramanian et al 25) and Buko et al, 26) in which they reported that modification of synaptosomal membranes 25) and liver plasma membranes 26) with HNE caused an increased lipid fluidity of the membranes.…”

Proteomic Analysis of Protein Expression and Oxidative Modification in R6/2 Transgenic Mice

“…Oxidatively modified proteins are generally dysfunctional, losing catalytic or structural integrity (29,51,52). Oxidative damage to proteins is of importance in explaining metabolic dysfunctions associated to oxidative stress-mediated diseases.…”

Redox Proteomics in Selected Neurodegenerative Disorders: From Its Infancy to Future Applications

“…Through hydration, the C-O bond breaks, resulting in 4-hydroxy-2-nonenal. As noted, once formed, HNE can covalently attach to proteins by Michael addition, which alters protein structure (361) and causes a loss of protein function and activity (139).…”

“…Protein-bound HNE in brain and progression of Alzheimer's disease. As just discussed, HNE is a reactive product of lipid peroxidation, and this a,b-unsaturated alkenal binds to Cys, His, or Lys residues of proteins, thereby changing the conformation and function of proteins (66,79,139,187,361). In AD, HNE has been found to be significantly elevated in AD brain, plasma, and CSF (187,238,297,348).…”