Keratin-1 Interaction with C-reactive Protein and Oxidized Low Density Lipoprotein

• Main Authors: HSU, PI-CHIN, 許碧芹
• Other Authors: HUANG, KUANG-TSE
• Format: Others
• Language: zh-TW
• Published: 2016
• Online Access: http://ndltd.ncl.edu.tw/handle/62794966123162674991

碩士 === 國立中正大學 === 化學工程研究所 === 104 === Atherosclerosis is considered as a chronic inflammatory disease. C-reactive protein (CRP) is a sensitive indicator of inflammation and plays an important role in atherosclerosis. Oxidized low density lipoprotein (oxLDL) contributes to the atherosclerotic plaque formation by induction of endothelial cell activation and dysfunction, foam cell formation, and smooth muscle cell proliferation. Previously, we found that keratin-1 (KRT1) binds to CRP and interfered the CRP effect on reduction of nitric oxide release. To further elucidate the interaction of KRT1 with CRP and oxLDL, KRT1 gene was amplified from pcDNA3.1(+)-KRT1 using polymerase chain reaction (PCR) and inserted into pET29c with a 6X His-tag at the C-terminus, forming plasmid pET29-KRT1. pET29-KRT1 was transformed into E. coli. The KRT1 expression was induced by IsoPropyl-Beta-D-ThioGalactoside (IPTG) and purified using Ni2+-column. The interaction of KRT1 with CRP and oxLDL was assayed by the enzyme-linked immunosorbent assay (ELISA). Our results showed that KRT1 bound to CRP in the presence but not absence of Ca2+, suggesting that the binding of CRP to KRT1 may be through the B-face of CRP. We also confirmed that KRT1 bound to oxLDL and the binding affinity was dependent on the oxidation degree of oxLDL. The binding of KRT1 to CRP and oxLDL strongly suggests the involvement of KRT1 in the process of atherosclerosis.