Solubilization of proteins in dimethyl sulfoxide by permethylation: application to structural studies of apolipoprotein B.
The delipidated protein moiety, apolipoprotein B, of human low-density lipoproteins was permethylated in potassium butoxide/dimethyl sulfoxide with methyl iodide. The derivatized protein was soluble in dimethyl sulfoxide and, in the presence of sodium dodecyl sulfate, in an aqueous buffer. Analysis of the methylated apolipoprotein B by polyacrylamide gradient gel electrophoresis in the presence of sodium dodecyl sulfate revealed five discrete bands of lower molecular mass than that of the parent 265-kDa protein, which disappeared upon permethylation. The electrophoretic behavior of the methylated apolipoprotein B was distinctly different from that of the other methylated proteins studied, including transferrin, bovine serum albumin, aldolase, beta-lactoglobulin, and apolipoprotein A-I, all of which had a higher apparent molecular weight after permethylation as compared to the corresponding native polypeptide. Calculated on the basis of methylated standard proteins the five polypeptides of apolipoprotein B have apparent molecular masses of 9.0, 16.6, 25.6, 35.7, and 46.7 kDa. The results suggest that the protein moiety of human low-density lipoprotein consists of subunits. In general, the results indicate that the permethylation method can be used to solubilize hydrophobic proteins in organic solvents for structural studies.
D001055:Apolipoproteins B / Q000302:isolation & purification; D002850:Chromatography, Gel; D004121:Dimethyl Sulfoxide; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008077:Lipoproteins, LDL / Q000302:isolation & purification; D008745:Methylation; D012995:Solubility
D001055:Apolipoproteins B; D008077:Lipoproteins, LDL; D004121:Dimethyl Sulfoxide