Molecular cloning and expression of partial cDNAs and deduced amino acid sequence of a carboxyl-terminal fragment of human apolipoprotein B-100.
Apolipoprotein (apo) B-100 cDNAs were identified in a human liver cDNA library cloned in the expression vector lambda gt11. The beta-galactosidase-apoB-100 fusion protein was detected by two independently produced low density lipoprotein polyclonal antisera and by three apoB-100 monoclonal antibodies that crossreact with apoB-74. It was not recognized by two apoB-100 monoclonal antibodies that crossreact with apoB-26. The longest clone, lambda B8, was completely sequenced. It contains a 2.8-kilobase DNA fragment containing the codons for the carboxyl-terminal 836 amino acid residues of apo-B-100, as well as the 3' untranslated region of apoB-100 mRNA. We have thus mapped apoB-74 to the carboxyl-terminal portion of apoB-100. The deduced amino acid sequence of the cloned DNA matches the sequences of 14 apoB-100 peptides determined in our laboratory. Minor differences in amino acid sequence were noted in three of the peptides, suggesting polymorphism of apoB-100 at the protein and DNA levels. Secondary structure predictions reveal an unusual pattern for apolipoproteins, consisting of beta-structure (24%), alpha-helical content (33%), and random structure (30%). Ten amphipathic helical regions of 10-24 residues were identified. This carboxyl-terminal fragment of apoB-100 is considerably more hydrophobic than other apolipoproteins with known structure. Its lipid binding regions might include stretches of highly hydrophobic beta-sheets as well as amphipathic helices. Our findings on apoB structure might be important for understanding the role of apoB-100-containing lipoproteins in atherosclerosis.
Wei
C F
CF
Chen
S H
SH
Yang
C Y
CY
Marcel
Y L
YL
Milne
R W
RW
Li
W H
WH
Sparrow
J T
JT
Gotto
A M
AM
Chan
L
L
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D010582:Bacteriophage lambda / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D004274:DNA, Recombinant / Q000378:metabolism; D008077:Lipoproteins, LDL / Q000276:immunology; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B; D004274:DNA, Recombinant; D008077:Lipoproteins, LDL; D011994:Recombinant Proteins; D004247:DNA