| Regional Biophysics Meeting 2005, March 16-20, Zreče, Slovenia | [MembBiophys] |
Low density lipoprotein (LDL) still represents a research challenge with respect to the understanding of both the extremely complex structural organization of the particle and its precise role in the development of the pathological process of atherosclerosis. Various spectroscopic approaches are being used with the single idea to provide the information at the atomic resolution about this protein-lipid assembly. In this context our study aims to provide experimental data on the noncovalent interaction of the protein part (apo B with the molecular weight of cca. 500 kDa) with the lipid matrix (more than 3000 molecules). The approach is based on the combination of 1H NMR (600 MHz) spectroscopy with the X-band CW-EPR and thiol specific spin labeling. In the 1H spectra of LDL we concentrate on the composite spectral peak centered around 3.25 ppm which has been assigned to the methyl headgroups of phosphatidylcholine and sphingomyelin of the particle [Biochemistry 39(32) (2000) 9763]. We explore their surface solvation via relaxation enhancement using chromium oxalate as a polar paramagnetic relaxation compound. In addition, we selectively label cysteine residues in apo B of LDL by covalently attaching methanethiosulphonate spin label to the free thiol groups. This approach offers not only a direct method to analyze lipid-protein interaction by NMR but also identifies which region in the primary sequence of the protein is in contact with the certain type of lipid molecules. Moreover, the spin labeled LDL is further studied by EPR.
Email: kveder@irb.hr
Address: Ruđer Bošković Institute, Bijenička 54, 10000 Zagreb, Croatia