Submitted on August 20, 2007
Revised on November 15, 2007
Accepted on November 17, 2007
Proteomics and lipid composition of lipoproteins isolated at low salt concentrations in D2O/sucrose or in KBr
Marcus Ståhlman, Pia Davidsson, Ida Kanmert, Birgitta Rosengren, Jan Borén, Björn Fagerberg, and Germán camejo
AstraZeneca, Discovery, Molndal S-431 83
Corresponding Author: german.camejo{at}astrazeneca.com
Abstract: There is much interest on the significance of apolipoproteins and proteins that are non-covalently associated with lipoproteins. It is possible that the high ionic strength used for isolation of lipoproteins with KBr and NaI could alter the pattern of associated exchangeable proteins. Here we describe lipoprotein classes fractionation from up to 0.5 ml of serum or plasma with buffers of physiological ionic strength and pH prepared with deuterium oxide (D2O) and sucrose. An advantage of the D2O/sucrose procedure was that the lipoproteins could be directly analyzed by the techniques described without need for desalting. We compared the isolated lipoproteins with those obtained using ultracentrifugation in KBr from the same plasma pool. Electrophoretic homogeneity of the lipoproteins was very similar with the two methods as well as their lipid composition evaluated by HPLC. Two dimensional electrophoresis and surface-enhanced laser adsorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) indicated that the patterns of exchangeable proteins of VLDL isolated using with the two procedures were very similar. However, significant differences were found in the profiles of LDL and HDL indicating that the D2O/sucrose method allowed a more complete characterization of its exchangeable apolipoproteins and proteins.
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