Construction of SDF-1P2G54, a specific antagonist of CXCR4 / 南方医科大学学报
Journal of Southern Medical University
; (12): 55-60, 2012.
Article
en Zh
| WPRIM
| ID: wpr-265697
Biblioteca responsable:
WPRO
ABSTRACT
<p><b>OBJECTIVE</b>To obtain a specific antagonist of CXCR4, SDF-1P2G54 by mutating SDF-1 second proline (P) into glycin (G) and removing the α-helix of its C-terminal.</p><p><b>METHODS</b>SDF-1p2g54 gene amplified by PCR was inserted into the vector pET-30a (+) and transformed into Escherichia coli (E. coli) strain BL21. After IPTG induction of E. coli, the expressed recombinant protein was purified with nickel-affinity chromatography column under denaturing conditions and refolded with gradient dilution and ultra-filtration. The chemotactic effect of SDF-1P2G54 on Jurkat cells and its antagonistic effect against SDF-1 were determined by transwell assay; flow cytometry was used to assay the ability of SDF-1P2G54 to induce calcium influx and CXCR4 internalization in MOLT4 cells.</p><p><b>RESULTS</b>The recombinant protein SDF-1P2G54 completely lost the functions to activate CXCR4 or to induce transmembrane migration of Jurkat cells and calcium influx in MOLT4 cells, but maintained a high affinity to CXCR4. SDF-1P2G54 effectively inhibited the chemotactic effect of wild-type SDF-1 to Jurkat cells, and induced rapid CXCR4 internalization in MOLT4 cells.</p><p><b>CONCLUSION</b>SDF-1P2G54 is a new antagonist of CXCR4 with a potential value as an effective inhibitor of HIV-1 infection, cancer metastasis or other major diseases.</p>
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1
Base de datos:
WPRIM
Asunto principal:
Proteínas Recombinantes
/
Línea Celular
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Receptores CXCR4
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Quimiocinas CXC
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Escherichia coli
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Proteínas Mutantes
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Genética
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Metabolismo
Límite:
Humans
Idioma:
Zh
Revista:
Journal of Southern Medical University
Año:
2012
Tipo del documento:
Article