Faculty Collaboration Database

Spin-labeling study of the oxidative damage to low-density lipoprotein. Arch Biochem Biophys 1995 Jun 20;320(1):155-61

Date

06/20/1995

Pubmed ID

7793976

DOI

10.1006/abbi.1995.1354

Scopus ID

2-s2.0-0029013080 (requires institutional sign-in at Scopus site)   23 Citations

Abstract

In this study, we have spin-labeled the lysine and cysteine residues of low-density lipoprotein (LDL) using N-4-(2,2,6,6-tetramethylpiperidinyl-1-oxyl-4-yl) maleimide (MAL-6) and succinimidyl-2,2,5,5-tetramethyl-3-pyrroline-1-oxyl-3-carboxylate (SSL), respectively. The electron spin resonance (ESR) spectrum of SSL bound to LDL indicated that the nitroxide moiety was relatively mobile. In contrast, the ESR spectrum of MAL-6 bound to LDL showed that the nitroxide moiety was rotationally restricted. Using the continuous-wave power saturation technique in the presence of hydrophobic and hydrophilic paramagnetic relaxing agents, we have determined that (i) approximately 60-70% of lysine-bound SSL is exposed to the aqueous phase, (ii) approximately 30-40% of SSL-LDL is buried in a hydrophobic region, and (iii) MAL-6 bound to LDL is localized predominantly in the hydrophobic region. During Cu(2+)-initiated oxidation of spin-labeled LDL, nitroxide labels located in a hydrophobic environment were predominantly degraded. Nitroxide destruction was inhibited by butylated hydroxytoluene, indicating the role of lipid peroxidation in this process. ESR data also showed that Cu2+ binding to lysine is essential for LDL oxidation. The spin label methodology may be useful for the investigation of site-specific radical reactions in LDL.

Author List

Singh RJ, Feix JB, Mchaourab HS, Hogg N, Kalyanaraman B

Authors

Jimmy B. Feix PhD Professor in the Biophysics department at Medical College of Wisconsin
Neil Hogg PhD Associate Dean, Professor in the Biophysics department at Medical College of Wisconsin
Balaraman Kalyanaraman PhD Professor in the Biophysics department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Apolipoproteins B
Binding Sites
Cyclic N-Oxides
Cysteine
Electron Spin Resonance Spectroscopy
Free Radicals
Humans
In Vitro Techniques
Lipid Peroxidation
Lipoproteins, LDL
Lysine
Microwaves
Molecular Structure
Oxidation-Reduction
Spin Labels