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Identification and characterisation of variation in the human apolipoprotein B gene

Dunning, Alison Margaret; (1994) Identification and characterisation of variation in the human apolipoprotein B gene. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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This thesis describes the search for genetic variants of the human apolipoprotein B (apoB) gene that have the potential to affect serum lipid levels. ApoB has a central role in lipid metabolism, both in maintaining the structure of the LDL particle and acting as the ligand for the LDL-receptor. The apoB gene is a candidate gene for the development of coronary artery disease. Eight patients whose LDL had reduced binding to the LDL-receptor were screened for mutations in the gene region encoding the putative receptor binding domain (residues 3130 to 3630). None were found, and it was concluded that other regions of the apoB protein may also be involved in forming the ligand for the LDL-receptor. There is some evidence that regions of apoB encoded by exon 29 of the gene may also affect binding of LDL to the receptor and so this exon was also screened for variations. A common polymorphism was discovered in exon 29 of the gene which results in the substitution of serine for asparagine at residue 4311 of the mature protein. In population samples from four different ethnic groups this polymorphism is in complete linkage disequilibrium with a reported polymorphism in exon 26 of the same gene. In addition, these two polymorphisms show complete association with the Antigen group (x/y) protein polymorphism of apoB and thus these amino acid substitutions are candidates for being the molecular bases of the Ag(x/y) epitopes. In a sample of healthy Swedish males there was a trend for the allele encoding Leu2712, Ser4311 and Ag(x) to be associated with reduced mean serum levels of apoB and LDL-cholesterol and raised levels of HDL-cholesterol, although only the latter reached statistical significance in the population sample studied. The high degree of linkage disequilibrium existing across the entire apoB gene enabled a phylogenetic tree for the human apoB gene to be postulated. This parsimonious model assumed that each haplotype in the tree was derived from the preceding one by single base substitutions and one small deletion without obligate recombination events. The validity of this model was tested in a sample of Swedish myocardial infarction patients and age-matched controls. All the postulated haplotypes were seen unequivocally, however other rare, unpredicted haplotypes were also present and these occurred significantly more frequently in the patient than in the control group. Recombination events could explain the generation of the unpredicted haplotypes. Two of the unpredicted haplotypes, uniquely defined by the presence of the XbaI X-allele in combination with the allele encoding Ag(a), were seen unequivocally and were found to be associated with raised mean serum apoB levels. This reached statistical significance in the patient group. A search was made for other amino acid substitutions encoded by exon 29 of the apoB gene in a group of individuals with aberrant binding of LDL to the receptor. Only one was found, this resulted in the substitution of threonine for arginine at residue 4243 of mature apoB. The entire exon 29 of the human apoB gene was cloned and expressed in a baculovirus expression system. A peptide of approximately 60kDa was detected using the antibody Bsol16 on a western blot. This expressed peptide has the potential to be used to test whether the apoB region encoded by exon 29 has affinity for lipid and the LDL-receptor.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Identification and characterisation of variation in the human apolipoprotein B gene
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest.
URI: https://discovery.ucl.ac.uk/id/eprint/10102719
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