eprintid: 10107109
rev_number: 8
eprint_status: archive
userid: 695
dir: disk0/10/10/71/09
datestamp: 2020-08-06 11:32:01
lastmod: 2020-08-06 11:32:01
status_changed: 2020-08-06 11:32:01
type: thesis
metadata_visibility: show
creators_name: Liu, Haiqun
title: Expression and structure-functional studies of human apolipoproteinciii
ispublished: unpub
keywords: Biological sciences; Recombinant apoCIII proteins
note: Thesis digitised by ProQuest.
abstract: Apolipoprotein (Apo) CIII plays a key role in triglyceride (TG)-rich lipoprotein metabolism and is a risk factor for coronary heart disease (CHD). The study involved in this thesis is the first in vitro structure-functional study using recombinant apoCIII proteins. The physicochemical properties of recombinant wild type and A23T, a naturally occurring mutation that is associated with apoCIII deficiency and lower plasma TG levels, as well as three site-directed mutants of apoCIII, designed by molecular modelling and implicated in lipid binding (L9T/T20L, F64A/W65A) or lipoprotein lipase (LPL) inhibition (K21A), were compared. Relative lipid binding efficiencies of each apoCIII variants to 1.2-dimyristoyl-sn-glycero-3- phosphatidylcholine (DMPC) were: L9T/T20L>WT>K21A>A23T>F64A/W65A with an inverse correlation with size of the discoidal complexes formed. Physicochemical analysis (Trp fluorescence, circular dichroism (CD) and GdnHC1 denaturation) suggested that the stability of the resulting apoCIII:DMPC complexes were dependent on their lipid binding properties. The displacement of apoE by apoCIII variants were tested by gel filtration of apoE:dipalmitoylphosphatidylcholine (DPPC) discoidal complexes mixed with the various apoCIII variants. All apoCIII proteins bound to the apoE:DPPC complexes and the capacity to displace apoE from the complex was dependent on their lipid binding affinity. All the recombinant apoCIII proteins inhibited LPL in the presence or in the absence of apoCII, with F64A/W65A displaying the most inhibition, suggesting that apoCIII inhibition of LPL is independent of lipid binding and therefore due to the protein:protein interaction with apoCII and/or LPL. Taken together, our data suggest that the hydrophobic residues F64 and W65 are crucial for the lipid binding properties of apoCIII and the redistribution of the N- terminal helix of apoCIII (L9T/T20L permutation) can enhance the lipid binding properties of the protein. Additionally, the reduced lipid binding capacity of the naturally occurring mutation A23T could lead to reduced plasma apoCIII and lower plasma TG levels in carriers.
date: 2000
oa_status: green
full_text_type: other
thesis_class: doctoral_open
thesis_award: Ph.D
language: eng
thesis_view: UCL_Thesis
primo: open
primo_central: open_green
verified: verified_manual
full_text_status: public
pages: 224
institution: UCL (University College London)
department: Medicine
thesis_type: Doctoral
citation:        Liu, Haiqun;      (2000)    Expression and structure-functional studies of human apolipoproteinciii.                   Doctoral thesis  (Ph.D), UCL (University College London).     Green open access   
 
document_url: https://discovery.ucl.ac.uk/id/eprint/10107109/1/Expression_and_structure-funct.pdf