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The expression of recombinant human LOX-1 and identifying its mimic ligands by fluorescence polarization-based high throughput screening
Zhang T.; Huang Z.; Dai Y.; Chen X.; Zhu P.; Du G.
2006-10-01
Source PublicationJournal of Biotechnology
ISSN01681656
Volume125Issue:4Pages:492-502
Abstract

Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) was identified as a major receptor for oxidized low-density lipoprotein (oxLDL) in endothelial cells. LOX-1 critically mediates the endothelial dysfunction and the progression of atherosclerosis by oxLDL stimulation. It might be an important target for vascular endothelium. In order to obtain human LOX-1 and identify its mimic ligand for facilitating the study of LOX-1 function, a recombinant plasmid pPIC9K-His-hLOX-1 was structured and expressed human LOX-1 in Pichia pastoris GS115. Western blot analysis ensured the expressed recombinant human LOX-1 protein and a receptor-ligand binding assay showed that it had high binding affinity with oxLDL. With this receptor protein, a competitive fluorescence polarization-based high throughput screening method was established in a 384-well microplate to isolate the mimic ligands of human LOX-1. The evaluating parameter Z′ value of 0.72 for this method showed that fluorescence polarization-based high throughput screening assay was robust and the results had a high reliability. By the fluorescence polarization-based high throughput screening assay, a total of 20,316 chemicals were screened, and 2 chemicals were identified that they have a high affinity with human LOX-1. Competitive uptake DiI-oxLDL assay by human LOX-1 transfected CHO-K1 cells further confirmed that two chemicals block the uptake of DiI-oxLDL. And the preliminary results indicated that isolated mimic ligands may act as a function of antagonist. The discovery of human LOX-1 mimic ligand would benefit to further study the function of LOX-1 and identify a novel avenue for prevention and treatment atherosclerosis. © 2006 Elsevier B.V. All rights reserved.

KeywordAtherosclerosis Endothelial Dysfunction Fluorescence Polarization High Throughput Screening Lectin-like Low-density Lipoprotein Receptor-1
DOI10.1016/j.jbiotec.2006.03.040
URLView the original
Indexed BySCI
Language英语
WOS Research AreaBiotechnology & Applied Microbiology
WOS SubjectBiotechnology & Applied Microbiology
WOS IDWOS:000241003200005
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Cited Times [WOS]:6   [WOS Record]     [Related Records in WOS]
Document TypeJournal article
CollectionInstitute of Chinese Medical Sciences
AffiliationChinese Academy of Medical Sciences
Recommended Citation
GB/T 7714
Zhang T.,Huang Z.,Dai Y.,et al. The expression of recombinant human LOX-1 and identifying its mimic ligands by fluorescence polarization-based high throughput screening[J]. Journal of Biotechnology,2006,125(4):492-502.
APA Zhang T.,Huang Z.,Dai Y.,Chen X.,Zhu P.,&Du G..(2006).The expression of recombinant human LOX-1 and identifying its mimic ligands by fluorescence polarization-based high throughput screening.Journal of Biotechnology,125(4),492-502.
MLA Zhang T.,et al."The expression of recombinant human LOX-1 and identifying its mimic ligands by fluorescence polarization-based high throughput screening".Journal of Biotechnology 125.4(2006):492-502.
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