Molecular cloning of human LDL apolipoprotein B cDNA. Evidence for more than one gene per haploid genome.
We have isolated an apolipoprotein B (apo B) clone (pXB1) from a human liver cDNA expression library, by immunoselection with a polyclonal antibody to human low density lipoprotein. pXB1 was used to isolate 3 clones (pB2, pB3 and pB4), containing cDNA inserts spanning a region of 3.75 kbp, from a second human liver cDNA library. We report the sequence of 1359 nucleotides at the 3' end of the pB4 cDNA insert and the amino acids encoded by this sequence. The cDNA inserts of pBX1 and pB2 overlapped the sequenced portion of pB4. pB2 contained an EcoR1 restriction site (resulting in a Glu-Lys replacement) which is not present in pB3 or pB4 and pB3 contained an MspI site not present in pB2 or pB4. Since all 3 clones were derived from the mRNA of a single human liver, we suggest that the human haploid genome contains more than one functional apo B gene. Labelled probes spanning almost the whole of the pB4 cDNA insert hybridized with RNA from human liver and small intestine, showing that the apo B mRNAs from these two tissues have nucleotide sequences in common. The nucleotide sequence in human liver apo B mRNA is probably longer than 12 kb, showing that the MW of monomeric apo B is at least 350kd. Clone pB4 hybridized with mRNA of similar length in rabbit liver and small intestine. These results raise the possibility that the low MW apo B synthesized in the intestine (B-48) and the high MW apo B synthesized in the liver (B-100) are translated from the same mRNA. The expression products of fragments of pB4 cloned into an expression vector were blotted with monoclonal antibodies to human LDL. The results suggest that the cDNA insert in pB4 encodes a part of apo B common to B-48 and B-100 and a region close to the recognition site for the LDL receptor.
D000818:Animals; D001055:Apolipoproteins B / Q000235:genetics*; D001483:Base Sequence; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D005796:Genes; D005822:Genetic Vectors; D006801:Humans; D007421:Intestine, Small / Q000378:metabolism; D008077:Lipoproteins, LDL / Q000235:genetics*; D008099:Liver / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics
D001055:Apolipoproteins B; D008077:Lipoproteins, LDL; D012333:RNA, Messenger; D004247:DNA