PREPARATION OF APOLIPOPROTEIN OF VERY LOW DENSITY LIPOPROTEIN FROM EGG YOLK GRANULES

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Abstract
The very low density lipoproteins VLDL(MF) were isolated from the yolk granules of hen's egg by a combination of preparative ultracentrifugation and agarose gel filtration chromatography. Ultra‐centrifugal, electrophoretic and chromatographic analyses were performed to characterize and assess the purity of the VLDL(MF) and delipidated VLDL. Flotation velocity. analysis of VLDLCMF) indicated a heterogeneous preparation with two major floating boundaries with S°f, rates of 75s and 207S Alkylation of VLDL(MF) with iodoacetamide prevented aggregation of VLDL(MF) due to oxidation of sulfhydryl groups and decreased S°f rates to 41s and 108s. Concanavalin A affinity chromatography of VLDL(MF) produced a retained and an unretained fraction. The retained fraction contained twice as much total carbohydrates as the unfractionated VLDL(MF), while the unretained VLDL(MF) contained half as much carbohydrate. All fractions were glycoproteins containing hexose, hexosamine and sialic acid residues. VLDL(MF) was partially delipidated with n‐heptane which preferentially extracted the neutral lipids and preserved the solubility of the phospholipid‐protein residues. Two fractions of the partially delipidated VLDL(MF) were isolated by gel filtration and characterized by analytical ultracentrifugation and by phospholipid/protein ratios. VLDL(MF) was totally delipidated with sodium deoxycholate (NaDOC) and ethanol‐ether procedures. Removal of lipids by both methods produced aggregated apoproteins; however, the NaDOC apoprotein remained soluble, whereas that obtained from ethanol‐ether precipitated, but could be solubilized in sodium dodecylsulfate or urea.