High serum levels of low density lipoprotein (LDL) are associated with atherosclerosis, the leading cause of death in the USA. The long term goal of this proposal is to develop new treatments that lower LDL levels and alleviate cardiovascular disease. LDL is derived from a precursor particle called very low density lipoprotein (VLDL) which is assembled in and secreted from the liver. Soth LDL and VLDL are composed of fats, cholesterol, cholesterol esters, phospholipids and a single molecule of apolipoprotein S (ApoS). The amount of VLDL secreted from the liver in not regulated by changes in the level of ApoS expression which is relatively constant. Instead, in the absence of sufficient lipid, ApoS is degraded in the cell. This degradation pathway represents a target for lowering VLDL assembly and thereby decreasing LDL levels. The assembly of VLDL is thought to be a two-step process. In the first step that requires the ER resident protein microsomal triglyceride transfer protein (MTP), ApoS forms a dense 'initiation particle'that contains primarily phospholipids. In the second step the initiation particle fuses with a lipid droplet in the ER to form VLDL. The second step also requires MTP. How the initiation particle is formed and how MTP performs its functions are not known. To fill this gap this proposal will use a prokaryotic cell free translation system to produce initiation particles in vitro. This system will be used to test the role of lipids, MTP and the structure of ApoS in the formation of lipoprotein particles.
The specific aims are: 1) To express N-terminal fragments of ApoS in a cell free system supplemented with phospholipids and phospholipid-triacylglycerol emulsions to test the hypothesis that cotranslational interaction of ApoS with lipids is spontaneous and essential for the formation of the initiation complex. The results will reveal the role of successive domains of ApoS in lipid recruitment and the mechanism for the formation of the initiation complex. 2) To determine the role of MTP in the folding and lipid recruitment by the N-terminal domains of ApoS using the cell free system. The successful completion of this proposal will provide new mechanistic details about the VLDL assembly pathway that can be used to target drug discovery efforts to lower LDL levels by preventing its secretion from the cell. High serum levels of low density lipoprotein (LOL) are associated with atherosclerosis, one the leading causes of death in the USA. The long term goal of this proposal is to develop new treatments that lower LOL levels and alleviate cardiovascular disease. The research proposed will provide new mechanistic details about assembly of the precursor to LOL, very low density lipoprotein (VLOL), that can be used to target drug discovery efforts to lower LOL levels by preventing VLOL secretion from the cell.
High serum levels of low density lipoprotein (LDL) are associated with atherosclerosis, one the leading causes of death in the USA. The long term goal of this proposal is to develop new treatments that lower LDL levels and alleviate cardiovascular disease. The research proposed will provide new mechanistic details about assembly of the precursor to LDL, very low density lipoprotein (VLDL), that can be used to target drug discovery efforts to lower LDL levels by preventing VLDL secretion from the cell.
| Mitsche, Matthew A; Packer, Laura E; Brown, Jeffrey W et al. (2014) Surface tensiometry of apolipoprotein B domains at lipid interfaces suggests a new model for the initial steps in triglyceride-rich lipoprotein assembly. J Biol Chem 289:9000-12 |
| Miller, Sharon A; Burnett, John R; Leonis, Mike A et al. (2014) Novel missense MTTP gene mutations causing abetalipoproteinemia. Biochim Biophys Acta 1842:1548-54 |
| Mitsche, Matthew A; Wang, Libo; Jiang, Z Gordon et al. (2009) Interfacial properties of a complex multi-domain 490 amino acid peptide derived from apolipoprotein B (residues 292-782). Langmuir 25:2322-30 |
