Apolipoprotein B: removal of lipids by sodium cholate and reassociation of a lipid-free apoprotein with dipalmitoyl phosphatidylcholine.
Apolipoprotein B (apoB) of human plasma low-density lipoprotein has been solubilized with sodium cholate added in an amount highly above its critical micellar concentration. During isolation by gel exclusion chromatography on Sepharose CL-4B, the apoB forms mixed micelles of protein and detergent that are free of endogenous lipids. The circular dichroic spectra of the sodium cholate-solubilized apoB indicate significant heterogeneity within the fractions obtained by gel chromatography. The peak position fraction of apoB taken from the column was used for reassociation with dipalmitoyl phosphatidylcholine (DPPC). A soluble apoB-DPPC complex has been prepared by incubation of apoB-sodium cholate and DPPC-sodium cholate solutions at 42 degrees C, followed with detergent removal by extensive dialysis in the presence of a XAD-2 ion-exchange resin. Data from negative-stain electron microscopy suggests the incorporation of solubilized apoB into single-bilayer phospholipid vesicles. Upon reassociation with phospholipid, a shift (to shorter wavelengths) occurs in the intrinsic fluorescence of the apoB, thus indicating a transfer of tryptophan residues to a more hydrophobic environment. Sodium dodecylsulfate-polyacrylamide electrophoresis gives a single band (apparent Mr 370,000) for apoB after solubilization, purification and interaction with the phospholipid.
D001055:Apolipoproteins B / Q000097:blood* / Q000302:isolation & purification; D019826:Cholic Acid; D002793:Cholic Acids; D002942:Circular Dichroism; D006801:Humans; D008077:Lipoproteins, LDL / Q000097:blood / Q000302:isolation & purification; D008854:Microscopy, Electron; D011485:Protein Binding; D011487:Protein Conformation; D011663:Pulmonary Surfactants / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
D001055:Apolipoproteins B; D002793:Cholic Acids; D008077:Lipoproteins, LDL; D011663:Pulmonary Surfactants; D019826:Cholic Acid