Site-directed mutagenesis of an apolipoprotein E mutant, apo E5(Glu3----Lys) and its binding to low density lipoprotein receptors.
Apo E5(Glu3----Lys) is a naturally occurring apolipoprotein E (apo E) mutant found in patients with hyperlipoproteinemia and atherosclerosis. It has been shown to have a high affinity for low density lipoprotein (LDL) receptors. In this study, mutant apo E5 was produced by Chinese hamster ovary cells by means of an in vitro site-directed mutagenesis technique, and its LDL receptor binding activity was assessed. The apo E5 obtained from gene expression bound more readily to the LDL receptor than did plasma apo E3. The concentrations required for 50% competitive binding of 125I-labeled LDL to the LDL receptors were 58.9 ng/ml for plasma apo E3 and 25.7 ng/ml for the expressed apo E5. The expressed apo E5 displayed 229% normal binding. This result is highly consistent with that obtained with plasma apo E5, which showed 217% normal binding. Although the experimental apo E isoproteins contained more sialic acid than plasma apo E, the extent of sialylation had no effect on the receptor binding of apo E.
Dong
L M
LM
Department of Etiology and Pathophysiology, National Cardiovascular Center Research Institute, Osaka, Japan.
Yamamura
T
T
Tajima
S
S
Yamamoto
A
A
D000818:Animals; D001057:Apolipoproteins E / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D016466:CHO Cells / Q000378:metabolism; D006224:Cricetinae; D004247:DNA / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D007525:Isoelectric Focusing; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D011973:Receptors, LDL / Q000378:metabolism*; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics; D014162:Transfection
D001057:Apolipoproteins E; D011973:Receptors, LDL; D004247:DNA; D013762:Tetrahydrofolate Dehydrogenase