Refolding/Reconstitution of Apolipoprotein B and its Application

• Main Authors: Chu, Hsueh-Liang, 朱學亮
• Other Authors: Chang, Chia-Ching
• Format: Others
• Language: zh-TW
• Published: 2013
• Online Access: http://ndltd.ncl.edu.tw/handle/69773298726040212611

博士 === 國立交通大學 === 分子醫學與生物工程研究所 === 101 === Apolipoprotein (apo) B is the major structural component of very-low-density lipoprotein (VLDL) and low-density lipoprotein (LDL). The large size and extreme hydrophobicity of apoB render examination of its lipidation process an experimental challenge. In the present study, we showed that apoB lipidation and its folding reaction intermediates could be followed at single molecule level by a five-step (on-path) folding process. When carboxyl-terminal-truncated mutants apoB-29 and apoB-48, representing the amino-terminal 29% and 48%, respectively, of the full-length apoB, were used for comparison, we observed that the refolded apoB resembled the native LDL. Thus the process of lipidation in vitro recapitulates that of biosynthesis of apoB. Furthermore, hydrophobic super-paramagnetic iron oxide nanoparticles (SPIONPs) and the anti-cancer drug M4N (tetra-O-methyl nordihydroguaiaretic acid), used as an imaging enhancer and lipophilic drug model, respectively, were incorporated into the reconstituting apoB lipoparticle (rABL), leading to the formation of SPIONPs- and M4N- containing rABL (SPIO@rABL and M4N@rABL, respectively). SPIO@rABL presented significant hepatic contrast enhancement in T2-weighted magnetic resonance imaging in BALB/c mice, suggesting its potential application as a medical imaging contrast agent. M4N@rABL could reduce the viability of the cancer cell line A549. The current study also demonstrates that apoB can be used as hydrophobic/amphiphilic molecules carrier and be monitored at single molecule level, too.